KLK5 / 7+IL-4 targeted antibody and its use

Abstract

Aspects of this application provide bispecific antibodies and methods of using them for promoting barrier function and reducing inflammation to treat conditions such as Netherton syndrome, eosinophilic esophagitis, and atopic dermatitis. In some embodiments, the bispecific antibodies include at least one antigen-specific binding site that specifically binds to KLK5 and KLK7, and at least one antigen-specific binding site that specifically binds to IL-4 or its receptor. Certain aspects of this disclosure relate to the recognition that a loss of balance between endogenous KLK proteases and associated protease inhibitors can lead to barrier dysfunction, induce inflammation (as shown in Figure 1), and result in inflammatory conditions such as Netherton syndrome, eosinophilic esophagitis, and atopic dermatitis.

Description

[Technical Field]

[0001] Related applications This application claims priority under § 119(e) of the United States Patent Act to U.S. Provisional Application No. 63 / 578,959 filed on 25 August 2023, U.S. Provisional Application No. 63 / 549,274 filed on 2 February 2024, and U.S. Provisional Application No. 63 / 671,723 filed on 15 July 2024, the entire contents of each of these applications incorporated herein by reference.

[0002] Reference to electronic sequence listings The contents of the electronic sequence listing (A140770007WO00-SEQ-LJG.xml, size: 162,435 bytes, and creation date: August 22, 2024) are incorporated herein by reference in their entirety. [Background technology]

[0003] The skin, comprising tissue structures and cells such as epithelial cells and immune cells, together provides a functional and physical barrier that plays a crucial role in preventing the penetration of allergens into the body and responding to pathogens. Kallikrein (KLK) enzymes support skin homeostasis and wound healing by regulating desquamation and innate immunity. In healthy skin, the outermost layer of the epidermis is regularly shed through a KLK-driven proteolytic cascade, resulting in the degradation of corneodesmosomes and desquamation. KLK5 is understood to be the major activator of this proteolytic cascade. Autoactivating KLK5 enzymatically converts pro-KLK7 and pro-KLK14 into their active forms, stimulating a positive feedback loop that leads to the production of more pro-KLK5 via KLK14. These KLK enzymes are inhibited by endogenous serine protease inhibitors, such as lymphoepithelial Cazal-type-associated inhibitors. Dysregulation of KLK, including KLK5 and KLK7, is associated with skin disorders, inflammatory diseases, and cancer. For example, hyperactive kallikrein 5 and 7 are responsible for both hereditary and spontaneous epidermal barrier disorders (e.g., Netherton syndrome, eosinophilic esophagitis, atopic dermatitis). [Overview of the project] [Means for solving the problem]

[0004] Certain aspects of this disclosure relate to the recognition that a loss of balance between endogenous KLK proteases and associated protease inhibitors can lead to barrier dysfunction, induce inflammation (as shown in Figure 1), and result in inflammatory conditions such as Netherton syndrome, eosinophilic esophagitis, and atopic dermatitis. Furthermore, increased activity of T helper 2 (Th2) cytokines (e.g., IL-13 and IL-4) may further drive inflammation and contribute to barrier dysfunction and related conditions (as shown in Figure 2). In some embodiments, compositions and methods provided herein are useful for inhibiting KLK5 / KLK7 and IL-4 signaling to improve barrier function, reduce inflammation, and improve disease severity.

[0005] Furthermore, aspects of the present disclosure provide multispecific antibodies comprising at least one antigen-specific binding site that specifically binds to KLK5 and KLK7, and at least one antigen-specific binding site that specifically binds to IL-4 or its receptor. In certain embodiments, such multispecific antibodies are configured such that at least one arm binds to the active sites of KLK5 and KLK7, and at least another arm binds to IL-4 or the IL-4 receptor (IL-4R). In some embodiments, such multispecific antibodies are advantageous because they inhibit both KLK5 / 7 activity and IL-4 activity in a single molecule. In some embodiments, with respect to KLK5 / 7, such antibodies specifically bind to the active form of the enzyme, rather than to its proform.

[0006] In some embodiments, methods and related compositions, as well as antibodies for use in the methods, are provided that are useful for inhibiting KLK5 and KLK7 along with IL-4 activity, for the purpose of improving barrier function, reducing inflammation, and thereby improving disease severity. In particular, aspects of the present disclosure provide a bispecific antibody (referred to as an anti-KLK5 / KLK7+IL-4 targeted bispecific antibody) in which one arm comprises an antigen-specific binding site of a dual inhibitory antibody (referred to as an anti-KLK5 / KLK7 antibody) that has high binding affinity and specificity to the active sites of both KLK5 and KLK7, and the other arm specifically binds to IL-4 or its receptor and inhibits its activity. Accordingly, in some embodiments, the present disclosure provides methods and related antibody compositions for treating conditions associated with KLK5 and KLK7 dysregulation and / or abnormal IL-4 activity, such as Netherton syndrome, atopic dermatitis (with and without filaggrin mutations), eosinophilic esophagitis, prurigo nodularis, chronic pruritus of unknown cause (CPUO), asthma (e.g., KLK5-associated asthma), and ichthyosis vulgaris.

[0007] In some embodiments, the Disclosure provides a bispecific antibody (referred to as an anti-KLK5 / KLK7+IL-4 targeted bispecific antibody) comprising at least one antigen-specific binding site of a dual inhibitory antibody targeting KLK5 and KLK7, and at least one antigen-specific binding site that specifically binds to and inhibits IL-4. Accordingly, in some embodiments, the Disclosure provides methods and related antibody compositions for treating conditions associated with KLK5 and KLK7 dysregulation, in which Th2 cytokines, such as IL-4, also play a role in the pathogenesis of conditions such as Netherton syndrome, atopic dermatitis (with and without filaggrin mutations), eosinophilic esophagitis, prurigo nodularis, chronic pruritus of unknown cause (CPUO), asthma (e.g., KLK5-associated asthma), and ichthyosis vulgaris.

[0008] In some embodiments, the Disclosure provides a bispecific antibody comprising an antigen-specific binding site that specifically binds to KLK5 and KLK7, and an antigen-specific binding site that specifically binds to interleukin (IL)-4 or its receptor. In some embodiments, the antigen-specific binding sites that specifically bind to KLK5 and KLK7 bind to the active sites of KLK5 and KLK7 and inhibit the activity of the enzymes. In some embodiments, the antigen-specific binding sites that specifically bind to KLK5 and KLK7 include any one of the antibodies listed in Tables 1a and 1b: HC CDR1, HC CDR2, HC CDR3, LC CDR1, LC CDR2, and / or LC CDR3.

[0009] In some embodiments, the antigen-specific binding sites that specifically bind to KLK5 and KLK7 include HC CDR1, HC CDR2, and HC CDR3, which are heavy chain variable domains having the amino acid sequence of SEQ ID NO: 7, and LC CDR1, LC CDR2, and LC CDR3, which are light chain variable domains having the amino acid sequence of SEQ ID NO: 8. In some embodiments, the antigen-specific binding sites that specifically bind to KLK5 and KLK7 include HC CDR1, HC CDR2, and HC CDR3, which are heavy chain variable domains having the amino acid sequence of SEQ ID NO: 13, and LC CDR1, LC CDR2, and LC CDR3, which are light chain variable domains having the amino acid sequence of SEQ ID NO: 14. In some embodiments, the antigen-specific binding sites that specifically bind to KLK5 and KLK7 include HC CDR1, HC CDR2, and HC CDR3, which are heavy chain variable domains having the amino acid sequence of SEQ ID NO: 17, and LC CDR1, LC CDR2, and LC CDR3, which are light chain variable domains having the amino acid sequence of SEQ ID NO: 14. In some embodiments, the antigen-specific binding sites that specifically bind to KLK5 and KLK7 include heavy chain variable domains HC CDR1, HC CDR2, and HC CDR3 having the amino acid sequence of SEQ ID NO: 21, and light chain variable domains LC CDR1, LC CDR2, and LC CDR3 having the amino acid sequence of SEQ ID NO: 14.

[0010] In some embodiments, the antigen-specific binding sites that specifically bind to KLK5 and KLK7 include HC CDR1 having the amino acid sequence of SEQ ID NO: 1, HC CDR2 having the amino acid sequence of SEQ ID NO: 2, HC CDR3 having the amino acid sequence of SEQ ID NO: 3, LC CDR1 having the amino acid sequence of SEQ ID NO: 4, LC CDR2 having the amino acid sequence of SEQ ID NO: 5, and LC CDR3 having the amino acid sequence of SEQ ID NO: 6. In some embodiments, the antigen-specific binding sites that specifically bind to KLK5 and KLK7 include HC CDR1 having the amino acid sequence of SEQ ID NO: 9, HC CDR2 having the amino acid sequence of SEQ ID NO: 10, HC CDR3 having the amino acid sequence of SEQ ID NO: 11, LC CDR1 having the amino acid sequence of SEQ ID NO: 4, LC CDR2 having the amino acid sequence of SEQ ID NO: 5, and LC CDR3 having the amino acid sequence of SEQ ID NO: 12. In some embodiments, the antigen-specific binding sites that specifically bind to KLK5 and KLK7 include HC CDR1 having the amino acid sequence of SEQ ID NO: 9, HC CDR2 having the amino acid sequence of SEQ ID NO: 15, HC CDR3 having the amino acid sequence of SEQ ID NO: 16, LC CDR1 having the amino acid sequence of SEQ ID NO: 4, LC CDR2 having the amino acid sequence of SEQ ID NO: 5, and LC CDR3 having the amino acid sequence of SEQ ID NO: 12. In some embodiments, the antigen-specific binding sites that specifically bind to KLK5 and KLK7 include HC CDR1 having the amino acid sequence of SEQ ID NO: 18, HC CDR2 having the amino acid sequence of SEQ ID NO: 19, HC CDR3 having the amino acid sequence of SEQ ID NO: 20, LC CDR1 having the amino acid sequence of SEQ ID NO: 4, LC CDR2 having the amino acid sequence of SEQ ID NO: 5, and LC CDR3 having the amino acid sequence of SEQ ID NO: 12.

[0011] In some embodiments, the antigen-specific binding sites that specifically bind to KLK5 and KLK7 are the VH and / or VL sites of any one of the antibodies listed in Table 1a.

[0012] In some embodiments, the antigen - specific binding site that specifically binds to KLK5 and KLK7 comprises a VH comprising the amino acid sequence of SEQ ID NO: 7 and a VL comprising the amino acid sequence of SEQ ID NO: 8. In some embodiments, the antigen - specific binding site that specifically binds to KLK5 and KLK7 comprises a VH comprising the amino acid sequence of SEQ ID NO: 13 and a VL comprising the amino acid sequence of SEQ ID NO: 14. In some embodiments, the antigen - specific binding site that specifically binds to KLK5 and KLK7 comprises a VH comprising the amino acid sequence of SEQ ID NO: 17 and a VL comprising the amino acid sequence of SEQ ID NO: 14. In some embodiments, the antigen - specific binding site that specifically binds to KLK5 and KLK7 comprises a VH comprising the amino acid sequence of SEQ ID NO: 21 and a VL comprising the amino acid sequence of SEQ ID NO: 14.

[0013] In some embodiments, the antigen - specific binding site that specifically binds to IL - 4 or its receptor comprises any one of the HC CDR1, HC CDR2, HC CDR3, LC CDR1, LC CDR2, and / or LC CDR3 of the antibodies listed in Table 2.

[0014] In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R includes HC CDR1, HC CDR2, and HC CDR3, which are heavy chain variable domains having the amino acid sequence of SEQ ID NO: 154, and LC CDR1, LC CDR2, and LC CDR3, which are light chain variable domains having the amino acid sequence of SEQ ID NO: 155. In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R includes HC CDR1, HC CDR2, and HC CDR3, which are heavy chain variable domains having the amino acid sequence of SEQ ID NO: 174, and LC CDR1, LC CDR2, and LC CDR3, which are light chain variable domains having the amino acid sequence of SEQ ID NO: 175. In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R includes HC CDR1, HC CDR2, and HC CDR3, which are heavy chain variable domains having the amino acid sequence of SEQ ID NO: 194, and LC CDR1, LC CDR2, and LC CDR3, which are light chain variable domains having the amino acid sequence of SEQ ID NO: 195. In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R includes HC CDR1, HC CDR2, and HC CDR3 of the heavy chain variable domain having the amino acid sequence of SEQ ID NO: 214, and LC CDR1, LC CDR2, and LC CDR3 of the light chain variable domain having the amino acid sequence of SEQ ID NO: 215. In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R includes HC CDR1, HC CDR2, and HC CDR3 of the heavy chain variable domain having the amino acid sequence of SEQ ID NO: 234, and LC CDR1, LC CDR2, and LC CDR3 of the light chain variable domain having the amino acid sequence of SEQ ID NO: 235. In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R includes HC CDR1, HC CDR2, and HC CDR3 of the heavy chain variable domain having the amino acid sequence of SEQ ID NO: 254, and LC CDR1, LC CDR2, and LC CDR3 of the light chain variable domain having the amino acid sequence of SEQ ID NO: 255.

[0015] In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R includes HC CDR1 having the amino acid sequence of SEQ ID NO: 156, HC CDR2 having the amino acid sequence of SEQ ID NO: 157, HC CDR3 having the amino acid sequence of SEQ ID NO: 158, LC CDR1 having the amino acid sequence of SEQ ID NO: 159, LC CDR2 having the amino acid sequence of SEQ ID NO: 160, and LC CDR3 having the amino acid sequence of SEQ ID NO: 161. In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R includes HC CDR1 having the amino acid sequence of SEQ ID NO: 162, HC CDR2 having the amino acid sequence of SEQ ID NO: 163, HC CDR3 having the amino acid sequence of SEQ ID NO: 164, LC CDR1 having the amino acid sequence of SEQ ID NO: 165, LC CDR2 having the amino acid sequence of LGS, and LC CDR3 having the amino acid sequence of SEQ ID NO: 167. In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R includes HC CDR1 having the amino acid sequence of SEQ ID NO: 168, HC CDR2 having the amino acid sequence of SEQ ID NO: 169, HC CDR3 having the amino acid sequence of SEQ ID NO: 170, LC CDR1 having the amino acid sequence of SEQ ID NO: 171, LC CDR2 having the amino acid sequence of SEQ ID NO: 172, and LC CDR3 having the amino acid sequence of SEQ ID NO: 173. In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R includes HC CDR1 having the amino acid sequence of SEQ ID NO: 176, HC CDR2 having the amino acid sequence of SEQ ID NO: 177, HC CDR3 having the amino acid sequence of SEQ ID NO: 178, LC CDR1 having the amino acid sequence of SEQ ID NO: 179, LC CDR2 having the amino acid sequence of SEQ ID NO: 180, and LC CDR3 having the amino acid sequence of SEQ ID NO: 181.In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R includes HC CDR1 having the amino acid sequence of SEQ ID NO: 182, HC CDR2 having the amino acid sequence of SEQ ID NO: 183, HC CDR3 having the amino acid sequence of SEQ ID NO: 184, LC CDR1 having the amino acid sequence of SEQ ID NO: 185, LC CDR2 having the amino acid sequence of AAS, and LC CDR3 having the amino acid sequence of SEQ ID NO: 187. In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R includes HC CDR1 having the amino acid sequence of SEQ ID NO: 188, HC CDR2 having the amino acid sequence of SEQ ID NO: 189, HC CDR3 having the amino acid sequence of SEQ ID NO: 190, LC CDR1 having the amino acid sequence of SEQ ID NO: 191, LC CDR2 having the amino acid sequence of SEQ ID NO: 192, and LC CDR3 having the amino acid sequence of SEQ ID NO: 193. In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R includes HC CDR1 having the amino acid sequence of SEQ ID NO: 196, HC CDR2 having the amino acid sequence of SEQ ID NO: 197, HC CDR3 having the amino acid sequence of SEQ ID NO: 198, LC CDR1 having the amino acid sequence of SEQ ID NO: 199, LC CDR2 having the amino acid sequence of SEQ ID NO: 200, and LC CDR3 having the amino acid sequence of SEQ ID NO: 201. In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R includes HC CDR1 having the amino acid sequence of SEQ ID NO: 202, HC CDR2 having the amino acid sequence of SEQ ID NO: 203, HC CDR3 having the amino acid sequence of SEQ ID NO: 204, LC CDR1 having the amino acid sequence of SEQ ID NO: 205, LC CDR2 having the amino acid sequence of SEQ ID NO: 206, and LC CDR3 having the amino acid sequence of SEQ ID NO: 207.In some embodiments, the antigen-specific binding sites that specifically bind to IL-4 or IL-4R include HC CDR1 having the amino acid sequence of SEQ ID NO: 208, HC CDR2 having the amino acid sequence of SEQ ID NO: 209, HC CDR3 having the amino acid sequence of SEQ ID NO: 210, LC CDR1 having the amino acid sequence of SEQ ID NO: 211, LC CDR2 having the amino acid sequence of GAS, and LC CDR3 having the amino acid sequence of SEQ ID NO: 213. In some embodiments, the antigen-specific binding sites that specifically bind to IL-4 or IL-4R include HC CDR1 having the amino acid sequence of SEQ ID NO: 216, HC CDR2 having the amino acid sequence of SEQ ID NO: 217, HC CDR3 having the amino acid sequence of SEQ ID NO: 218, LC CDR1 having the amino acid sequence of SEQ ID NO: 219, LC CDR2 having the amino acid sequence of SEQ ID NO: 220, and LC CDR3 having the amino acid sequence of SEQ ID NO: 221. In some embodiments, the antigen-specific binding sites that specifically bind to IL-4 or IL-4R include HC CDR1 having the amino acid sequence of SEQ ID NO: 222, HC CDR2 having the amino acid sequence of SEQ ID NO: 223, HC CDR3 having the amino acid sequence of SEQ ID NO: 224, LC CDR1 having the amino acid sequence of SEQ ID NO: 225, LC CDR2 having the amino acid sequence of YTS, and LC CDR3 having the amino acid sequence of SEQ ID NO: 227. In some embodiments, the antigen-specific binding sites that specifically bind to IL-4 or IL-4R include HC CDR1 having the amino acid sequence of SEQ ID NO: 228, HC CDR2 having the amino acid sequence of SEQ ID NO: 229, HC CDR3 having the amino acid sequence of SEQ ID NO: 230, LC CDR1 having the amino acid sequence of SEQ ID NO: 231, LC CDR2 having the amino acid sequence of SEQ ID NO: 232, and LC CDR3 having the amino acid sequence of SEQ ID NO: 233.In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R includes HC CDR1 having the amino acid sequence of SEQ ID NO: 236, HC CDR2 having the amino acid sequence of SEQ ID NO: 237, HC CDR3 having the amino acid sequence of SEQ ID NO: 238, LC CDR1 having the amino acid sequence of SEQ ID NO: 239, LC CDR2 having the amino acid sequence of SEQ ID NO: 240, and LC CDR3 having the amino acid sequence of SEQ ID NO: 241. In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R includes HC CDR1 having the amino acid sequence of SEQ ID NO: 242, HC CDR2 having the amino acid sequence of SEQ ID NO: 243, HC CDR3 having the amino acid sequence of SEQ ID NO: 244, LC CDR1 having the amino acid sequence of SEQ ID NO: 245, LC CDR2 having the amino acid sequence of SEQ ID NO: 246, and LC CDR3 having the amino acid sequence of SEQ ID NO: 247. In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R includes HC CDR1 having the amino acid sequence of SEQ ID NO: 248, HC CDR2 having the amino acid sequence of SEQ ID NO: 249, HC CDR3 having the amino acid sequence of SEQ ID NO: 250, LC CDR1 having the amino acid sequence of SEQ ID NO: 251, LC CDR2 having the amino acid sequence of SEQ ID NO: 252, and LC CDR3 having the amino acid sequence of SEQ ID NO: 253. In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R includes HC CDR1 having the amino acid sequence of SEQ ID NO: 256, HC CDR2 having the amino acid sequence of SEQ ID NO: 257, HC CDR3 having the amino acid sequence of SEQ ID NO: 258, LC CDR1 having the amino acid sequence of SEQ ID NO: 259, LC CDR2 having the amino acid sequence of SEQ ID NO: 260, and LC CDR3 having the amino acid sequence of SEQ ID NO: 261.In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R comprises HC CDR1 having the amino acid sequence of SEQ ID NO: 262, HC CDR2 having the amino acid sequence of SEQ ID NO: 263, HC CDR3 having the amino acid sequence of SEQ ID NO: 264, LC CDR1 having the amino acid sequence of SEQ ID NO: 265, LC CDR2 having the amino acid sequence of SEQ ID NO: 267, and LC CDR3 having the amino acid sequence of SEQ ID NO: 268. In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R comprises HC CDR1 having the amino acid sequence of SEQ ID NO: 269, HC CDR2 having the amino acid sequence of SEQ ID NO: 270, HC CDR3 having the amino acid sequence of SEQ ID NO: 271, LC CDR1 having the amino acid sequence of SEQ ID NO: 272, LC CDR2 having the amino acid sequence of SAS, and LC CDR3 having the amino acid sequence of SEQ ID NO: 274.

[0016] In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R comprises VH comprising the amino acid sequence of SEQ ID NO: 154 and VL comprising the amino acid sequence of SEQ ID NO: 155. In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R comprises VH comprising the amino acid sequence of SEQ ID NO: 174 and VL comprising the amino acid sequence of SEQ ID NO: 175. In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R comprises VH comprising the amino acid sequence of SEQ ID NO: 194 and VL comprising the amino acid sequence of SEQ ID NO: 195. In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R comprises VH comprising the amino acid sequence of SEQ ID NO: 214 and VL comprising the amino acid sequence of SEQ ID NO: 215. In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R comprises VH comprising the amino acid sequence of SEQ ID NO: 234 and VL comprising the amino acid sequence of SEQ ID NO: 235. In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or IL-4R comprises VH comprising the amino acid sequence of SEQ ID NO: 254 and VL comprising the amino acid sequence of SEQ ID NO: 255.

[0017] In some embodiments, the antigen-specific binding site that specifically binds to IL-4 or its receptor is the VH and / or VL of any one of the antibodies in Table 2.

[0018] In some embodiments, the bispecific antibodies provided herein retain a binding affinity to KLK5 and KLK7 that is 20% or less lower than that of an anti-KLK5 / KLK7 antibody containing the same CDR and / or VH / VL as the bispecific antibody. In some embodiments, the bispecific antibodies retain a binding affinity to IL-4 that is 20% or less lower than that of an anti-IL-4 antibody containing the same CDR and / or VH / VL as the bispecific antibody. In some embodiments, the bispecific antibodies retain at least 80% of the inhibitory activity against KLK5 and KLK7 compared to an anti-KLK5 / KLK7 antibody containing the same CDR and / or VH / VL as the bispecific antibody. In some embodiments, the bispecific antibodies retain at least 80% of the inhibitory activity against IL-4 signaling compared to an anti-IL-4 antibody containing the same CDR and / or VH / VL as the bispecific antibody.

[0019] In some embodiments, the Disclosure provides a composition comprising a bispecific antibody provided herein and an acceptable carrier. In some embodiments, the Disclosure provides a method comprising administering the antibody or composition provided herein to a subject. In some embodiments, the subject has a skin barrier defect.

[0020] In some embodiments, the Disclosure provides a method for treating a skin barrier defect, comprising administering an effective amount of a bispecific antibody or composition provided herein to a target. In some embodiments, the skin barrier defect is associated with Netherton syndrome, atopic dermatitis, eosinophilic esophagitis, prurigo nodosa, chronic pruritus of unknown cause (CPUO), dry skin, asthma (particularly KLK5), ichthyosis vulgaris, or itching or chronic itching.

[0021] In some embodiments, the subjects have atopic dermatitis. In some embodiments, administration of the antibody or composition provided herein reduces ear thickness by more than 30% compared to subjects administered with an anti-IL-4 antibody containing the same CDR and / or VH / VL as the bispecific antibody. In some embodiments, administration reduces ear thickness by more than 30% compared to subjects administered with an anti-IL-4 antibody containing the same CDR and / or VH / VL as the bispecific antibody. In some embodiments, administration reduces skin erythema / bleeding by more than 30% compared to subjects administered with an anti-IL-4 antibody containing the same CDR and / or VH / VL as the bispecific antibody. In some embodiments, administration reduces skin erythema / bleeding by more than 30% compared to subjects administered with an anti-IL-4 antibody containing the same CDR and / or VH / VL as the bispecific antibody. In some embodiments, administration reduces skin erythema / bleeding by more than 30% compared to subjects administered with an anti-IL-4 antibody containing the same CDR and / or VH / VL as the bispecific antibody. In some embodiments, administration reduces skin peeling / erosion by more than 30% compared to subjects administered with an anti-IL-4 antibody containing the same CDR and / or VH / VL as the bispecific antibody. In some embodiments, administration reduces skin scaling / dryness by more than 30% compared to subjects administered with an anti-IL-4 antibody containing the same CDR and / or VH / VL as the bispecific antibody. In some embodiments, administration reduces skin scaling / dryness by more than 30% compared to subjects administered with an anti-IL-4 antibody containing the same CDR and / or VH / VL as the bispecific antibody. In some embodiments, administration reduces skin edema by more than 30% compared to subjects administered with an anti-IL-4 antibody containing the same CDR and / or VH / VL as the bispecific antibody. In some embodiments, administration reduces skin edema by more than 30% compared to subjects administered with an anti-IL-4 antibody containing the same CDR and / or VH / VL as the bispecific antibody.

[0022] The preceding and other aspects, embodiments, actions, functions, features, and embodiments of this instruction can be better understood from the following description in conjunction with the accompanying drawings.

[0023] The accompanying drawings are incorporated herein and constitute part of this specification, illustrating certain embodiments and, together with the written description, are useful in providing non-limiting examples of certain aspects of the compositions and methods disclosed herein. [Brief explanation of the drawing]

[0024] [Figure 1] This chart shows abnormal protease activation (e.g., abnormal KLK5, KLK7, and KLK14) that can lead to skin barrier deficiency-related diseases. [Figure 2] This graph shows that in conditions associated with epidermal barrier dysfunction, increased KLK5 / KLK7 activity leads to Th2 cell activation, which in turn further increases KLK5 / KLK7 activity. [Figure 3] (Figure 3A) This graph shows that inhibiting KLK5 / KLK7 and Th2 signaling in an Nc / Nga model of atopic dermatitis has a more beneficial effect than targeting either pathway alone. It shows that mice treated with KLK5 / KLK7-dual-Ab4 in combination with an anti-IL-4R antibody have a further reduction in ear thickness compared to mice treated with KLK5 / KLK7-dual-Ab4 or anti-IL-4R antibody alone. (Figure 3B) This graph shows that inhibiting KLK5 / KLK7 and Th2 signaling in an Nc / Nga model of atopic dermatitis has a more beneficial effect than targeting either pathway alone. It shows that mice treated with KLK5 / KLK7-dual-Ab4 in combination with an anti-IL-4R antibody have a reduced clinical skin score compared to mice treated with KLK5 / KLK7-dual-Ab4 or anti-IL-4R antibody alone. [Modes for carrying out the invention]

[0025] This disclosure is at least in part based on the development of a bispecific antibody (referred to as an anti-KLK5 / KLK7+IL-4 bispecific antibody) comprising one arm containing an antigen-binding site of a dual inhibitory antibody and its variants targeting KLK5 and KLK7, and the other arm containing an antigen-binding site of an anti-IL4 or anti-IL4R antibody. These dual inhibitory antibodies target KLK5 and KLK7 via a common, distinct antigen-specific binding site. Such dual inhibitory antibodies have high binding affinity and specificity for KLK5 and KLK7 (anti-KLK5 / KLK7 antibody). In some embodiments, a bispecific antibody is provided in which one arm contains an antigen-specific binding site of a dual inhibitory antibody targeting KLK5 and KLK7, and the other arm specifically binds to and inhibits the activity of IL-4. Furthermore, methods for using anti-KLK5 / KLK7+IL-4 bispecific antibodies and their variants for research, diagnostic / detection, and therapeutic applications are also provided, as well as anti-KLK5 / KLK7+IL-4 bispecific antibodies for use in such methods.

[0026] The preceding and other aspects, embodiments, actions, functions, features, and embodiments of this instruction can be better understood from the following description in conjunction with the accompanying drawings.

[0027] I. Definition To administer: As used herein, the terms “to administer” or “to administer” mean to provide an antibody or a composition thereof to a subject in a physiologically and / or pharmacologically useful manner (for example, to treat a condition of the subject).

[0028] Affinity-matured antibodies: In this specification, the term “affinity-matured antibody” is used to refer to an antibody obtained by adding one or more modifications to one or more CDRs that result in improved affinity (e.g., KD, kd, or ka) of the antibody to a target antigen compared to an unmodified parent antibody. Exemplary affinity-matured antibodies may, in some embodiments, have nanomolar or even picomolar affinity to a target antigen. Various procedures are available for generating affinity-matured antibodies, including screening of combination antibody libraries prepared using biodisplay. For example, Marks et al., BioTechnology, 10:779-783 (1992) describes affinity maturation by shuffling of VH and VL domains. Random mutagenesis of CDRs and / or framework residues has been described by Barbas et al., Proc.Nat.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). Selective mutagenesis sites and selective mutations at contact sites or hypermutation sites with activity-enhancing amino acid residues have been described in U.S. Patent No. 6,914,128B1.

[0029] Antibody: As used herein, the term “antibody” refers to a polypeptide comprising at least one immunoglobulin variable domain containing at least one distinct antigen-specific binding site, or a portion of an immunoglobulin variable domain (a paratope or a portion thereof) containing at least one distinct antigen-specific binding site. In some embodiments, the antibody comprises at least one distinct antigen-specific binding site that specifically binds to the active site of an enzyme. In some embodiments, the antibody is an IL-4 targeted antibody comprising at least one antigen-specific binding site that specifically binds to IL-4 or its receptor. In some embodiments, the antibody is a full-length antibody. In some embodiments, the antibody is a chimeric antibody. In some embodiments, the antibody is a humanized antibody. However, in some embodiments, the antibody is a Fab fragment, an F(ab')2 fragment, an Fv fragment, or an scFv fragment. In some embodiments, the antibody is a multispecific antibody (e.g., a bispecific antibody). In some embodiments, the antibody is a nanobody derived from a camelid antibody or a nanobody derived from a shark antibody. In some embodiments, the antibody is a diabody. In some embodiments, the antibody comprises a framework having a human germline sequence. In another embodiment, the antibody comprises a heavy chain constant domain selected from the group consisting of IgG, IgG1, IgG2, IgG2A, IgG2B, IgG2C, IgG3, IgG4, IgA1, IgA2, IgD, IgM, and IgE constant domains. In some embodiments, the antibody comprises a heavy (H) chain variable region or domain (abbreviated herein as VH) and / or a light (L) chain variable region or domain (abbreviated herein as VL). In some embodiments, the antibody comprises a constant region. The immunoglobulin constant domain refers to the heavy chain constant domain or the light chain constant domain. The amino acid sequences of the human IgG heavy chain and light chain constant domain, as well as their functional mutations, are known. In some embodiments, the heavy chain constant domain of the immunoglobulin comprises an Fc region. With respect to the heavy chain, in some embodiments, the heavy chain of the antibody described herein may be an alpha (α), delta (Δ), epsilon (ε), gamma (γ), or mu (μ) heavy chain.In some embodiments, the heavy chains of the antibodies described herein may include human alpha (α), delta (Δ), epsilon (ε), gamma (γ), or mu (μ) heavy chains. In some embodiments, the antibodies described herein include human gamma-1 CH1, CH2, and / or CH3 domains. In some embodiments, the amino acid sequence of the antibody includes the amino acid sequence of the human gamma (γ) heavy chain constant region, such as those known in the art. Non-limiting examples of human constant region sequences are described in the art; see, for example, U.S. Patent No. 5,693,780 and Kabat EA et al., (1991) cited above. In some embodiments, the antibody includes a heavy chain having an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 98%, or at least 99% identical to any of the heavy chain constant regions provided herein. In some embodiments, the antibody comprises a light chain having an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 98%, or at least 99% identical to any of the light chain constant regions provided herein. In some embodiments, the antibody is modified, for example, by glycosylation, phosphorylation, SUMOylation, and / or methylation. In some embodiments, the antibody is a glycosylated antibody conjugated to one or more sugar or carbohydrate molecules. In some embodiments, one or more sugar or carbohydrate molecules are conjugated to the antibody via N-glycosylation, O-glycosylation, C-glycosylation, glyciation (GPI anchor attachment), and / or phosphoglycosylation. In some embodiments, one or more sugar or carbohydrate molecules are monosaccharides, disaccharides, oligosaccharides, or glycans. In some embodiments, one or more sugar or carbohydrate molecules are branched oligosaccharides or branched glycans. In some embodiments, one or more sugar or carbohydrate molecules include mannose units, glucose units, N-acetylglucosamine units, or phospholipid units. In some embodiments, the antibody is a construct comprising a polypeptide containing one or more antigen-binding fragments of the present disclosure linked to a linker polypeptide or an immunoglobulin constant domain.Linker polypeptides consist of two or more amino acid residues linked by peptide bonds and are used to link one or more antigen-binding moieties. Examples of linker polypeptides have been reported (see, for example, Holliger, P., et al. (1993) Proc. Natl. Acad. Sci. USA 90:6444-6448; Poljak, RJ, et al. (1994) Structure 2:1121-1123). Furthermore, antibodies may be part of larger immunoadhesion molecules formed by the covalent or noncovalent association of an antibody or antibody moiety with one or more other proteins or peptides. Examples of such immunoadhesion molecules include the use of streptavidin core regions to construct tetrameric scFv molecules (Kipriyanov, SM, et al. (1995) Human Antibodies and Hybridomas 6:93-101), and the use of cysteine ​​residues, marker peptides, and C-terminal polyhistidine tags to construct divalent biotinylated scFv molecules (Kipriyanov, SM, et al. (1994) Mol.Immunol.31:1047-1058).

[0030] Approximately: As used herein, the terms “approximately” or “about” refer to values ​​similar to the described reference values ​​when applied to the values ​​of one or more subjects. In certain embodiments, unless otherwise stated or evident from the content (except where such numbers may exceed 100% of the possible values), the terms “approximately” or “about” refer to a range of values ​​that fall within 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less in either direction (above or below) the described reference values.

[0031] Bispecific antibodies: As used herein, the term “bispecific antibodies” refers to an antibody containing two distinct antigen-specific binding sites, or, in combination, two antibodies (covalently or noncovalently) linked, each containing two distinct antigen-specific binding sites. Non-limiting examples of the format or structure of bispecific antibodies are provided in Labrijn, AF, et al., Bispecific antibodies: a mechanistic review of the pipeline, Nature Reviews Drug Discovery volume 18, pages 585-608 (2019) and Brinkmann U and Kontermann EE, The making of bispecific antibodies, MAbs. 2017 Feb / Mar;9(2):182-212, the entire contents of each of these are incorporated herein by reference.

[0032] CDR: As used herein, the term “CDR” refers to the complementarity-determining region within the antibody variable sequence. A typical antibody molecule usually contains a heavy chain variable region or heavy chain variable domain (VH) and a light chain variable region or light chain variable domain (VL) involved in antigen binding. The VH and VL regions can be further subdivided into hypervariable regions also known as “complementarity-determining regions” (“CDR”) and more conserved regions known as “framework regions” (“FR”). Each VH and VL typically consists of three CDRs and four FRs, arranged from the amino terminus to the carboxyl terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The scope of the framework regions and CDRs can be precisely identified using methodologies known in the art, such as the Kabat definition, IMGT definition, Chothia definition, AbM definition, and / or contact definition, all of which are well known in the art.For example, Kabat, EA, et al. (1991) Sequences of Proteins of Immunological Interest, Fifth Edition, USDepartment of Health and Human Services, NIH Publication No. 91-3242, IMGT (registered trademark), the international ImMunoGeneTics information system (registered trademark) http: / / www.imgt.org, Lefranc, M.-P. et al., Nucleic Acids Res.,27:209-212(1999), Ruiz,M.et al.,Nucleic Acids Res.,28:219-221(2000),Lefranc,M.-P.,Nucleic Acids Res.,29:207-209(2001),Lefranc,M.-P.,Nucleic Acids Res.,31:307-310(2003), Lefranc, M.-P.et. al.,In Silico Biol.,5,0006(2004)[[Epub]],5:45-60(2005),Lefranc,M.-P.et al.,Nucleic Acids Res.,33:D593-597(2005),Lefranc,M.-P.et al.,Nucleic Acids Res.,37:D1006-1012(2009), Lefranc,M.-P.et al.,Nucleic Acids Res.,43:D413-422(2015),Chothia et al.,(1989)Nature 342:877,Chothia,C.et al. (1987) J. Mol. Biol. 196:901-917, Al-lazikani et al. See al(1997)J.Molec.Biol.273:927-948 and Almagro,J.Mol.Recognit.17:132-143(2004). Also see hgmp.mrc.ac.uk and bioinf.org.uk / abs. As used herein, CDR may mean CDR as defined by any method known in the art.Two antibodies having the same CDR means that the amino acid sequences of the CDRs of the two antibodies are the same, as determined by the same method, for example, the IMGT definition.

[0033] In certain embodiments, three CDRs are present in each of the heavy and light chain variable regions, designated as CDR1, CDR2, and CDR3 with respect to each variable region. As used herein, the term “CDR set” refers to a group of three CDRs occurring in a single variable region capable of binding to an antigen. The exact boundaries of these CDRs are defined differently depending on the system. The system described by Kabat (Kabat et al., Sequences of Proteins of Immunological Interest (National Institutes of Health, Bethesda, Md. (1987) and (1991)) not only provides a clear residue numbering system applicable to any variable region of an antibody, but also provides precise residue boundaries that define three CDRs. These CDRs are sometimes called Kabat CDRs. The sub-parts of the CDRs may be designated as L1, L2, and L3, or H1, H2, and H3, where "L" and "H" designate the light chain region and heavy chain region, respectively. These regions are sometimes called Chothia CDRs and have boundaries that overlap with Kabat CDRs. Other boundaries that define CDRs that overlap with Kabat CDRs are described by Padlan (FASEB J.9:133-139 (1995)) and MacCallum (J Mol Biol This is described in 262(5):732-45(1996). Furthermore, other CDR boundary definitions may not strictly adhere to one of the above systems and, although they overlap with Kabat CDRs, they may be shortened or extended in consideration of predictions or experimental results that certain residues or groups of residues, or even the entire CDR, do not significantly affect antigen binding. The methods used herein may utilize CDRs defined according to any of these systems, but preferred embodiments use Kabat or Chothia-defined CDRs.

[0034] CDR-transplanted antibody: As used herein, the term "CDR-transplanted antibody" refers to an antibody that contains heavy chain variable region sequences and light chain variable region sequences derived from one species, but in which one or more sequences of the VH and / or VL CDR regions are replaced with CDR sequences from another species. For example, in an antibody having mouse heavy chain variable region and light chain variable region, one or more mouse CDRs (e.g., CDR3) are replaced with human CDR sequences.

[0035] Chimeric antibody: As used herein, the term "chimeric antibody" refers to an antibody that contains a heavy chain variable region sequence and a light chain variable region sequence derived from one species and a constant region sequence derived from another species, such as an antibody in which the mouse heavy chain variable region and light chain variable region are linked to the human constant region.

[0036] Complementary: As used herein, the term “complementary” refers to the ability of two nucleotides or two sets of nucleotides to form a precise pairing. Specifically, complementarity is a term that characterizes the degree of hydrogen bond pairing that results in a bond between two nucleotides or two sets of nucleotides. For example, if a base at one position of an oligonucleotide can form a hydrogen bond with a base at a corresponding position in a target nucleic acid (e.g., mRNA), then those bases are considered complementary to each other at that position. Base pairings can include both canonical Watson-Crick base pairings and non-Watson-Crick base pairings (e.g., Wobble base pairings and Hoogsteen base pairings). For example, in some embodiments, in the case of complementary base pairing, an adenosine-type base (A) is complementary to a thymidine-type base (T) or a uracil-type base (U), a cytosine-type base (C) is complementary to a guanosine-type base (G), and universal bases such as 3-nitropyrrole or 5-nitroindole can hybridize to any A, C, U, or T and are considered complementary to them. Inosine (I) is also considered a universal base in the art and is considered complementary to any of A, C, U, or T.

[0037] Conservative amino acid substitutions: As used herein, “conservative amino acid substitutions” refer to amino acid substitutions that do not alter the relative charge or size properties of the protein being substituted. Variants can be prepared according to methods for modifying polypeptide sequences known to those skilled in the art, for example, according to references that summarize such methods, e.g., Molecular Cloning: A Laboratory Manual, J. Sambrook, et al., eds., Fourth Edition, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, 2012, or according to methods found in Current Protocols in Molecular Biology, FMAusubel, et al., eds., John Wiley & Sons, Inc., New York. Conservative amino acid substitutions include substitutions made between amino acids in the following groups: (a) M, I, L, V; (b) F, Y, W; (c) K, R, H; (d) A, G; (e) S, T; (f) Q, N; and (g) E, D.

[0038] Cross-reactivity: As used herein, the term “cross-reactivity” refers to the property of a drug to specifically bind to multiple antigens of similar type or class (e.g., multiple homologous, paralogous, or orthologous antigens) with similar affinity or binding strength. For example, in some embodiments, an antibody that cross-reactive to human and non-human primate antigens of similar type or class (e.g., human KLK5 and non-human primate KLK5, human KLK7 and non-human primate KLK7) can bind to human and non-human primate antigens with similar affinity or binding strength. In some embodiments, the antibody cross-reactive to human and rodent antigens of similar type or class. In some embodiments, the antibody cross-reactive to rodent antigens and non-human primate antigens of similar type or class. In some embodiments, the antibody cross-reactive to human, non-human primate, and rodent antigens of similar type or class.

[0039] Dual-inhibitory antibody: As used herein, the term “dual-inhibitory antibody” refers to an antibody that targets at least two (e.g., 2, 3) different antigens via a common individual antigen-specific binding site and inhibits the activity of those antigens. In some embodiments, a dual-inhibitory antibody targets at least two different proteins (e.g., expressed from two different genes (e.g., endogenous genes, e.g., homologs, paralogs)) via a common individual antigen-specific binding site and inhibits the activity of at least two different proteins (e.g., enzymes such as proteases). In some embodiments, a dual-inhibitory antibody targets at least two different proteases (e.g., expressed from two different endogenous genes, e.g., KLK5 and KLK7) via a common individual antigen-specific binding site and inhibits the activity of at least two different proteases. In some embodiments, the common individual antigen-specific binding site binds to similar (e.g., homologous) domains shared between or among at least two different antigens. For example, in some embodiments, the common individual antigen-specific binding site binds to similar (e.g., homologous) catalytic domains or substrate-binding sites shared between or among at least two different enzymes, e.g., proteases. In some embodiments, the common individual antigen-specific binding site of a dual inhibitory antibody includes amino acids in one or more complementarity-determining regions of the antibody. In some embodiments, the common individual antigen-specific binding site of a dual inhibitory antibody is located within the heavy chain variable region and / or light chain variable region of the antibody. In some embodiments, the common individual antigen-specific binding site of a dual inhibitory antibody includes one or more complementarity-determining regions in the heavy chain variable region and / or light chain variable region of the antibody. In some embodiments, the common individual antigen-specific binding site of a dual inhibitory antibody includes HC CDR1, HC CDR2, HC CDR3, LC CDR1, LC CDR2, and LC CDR3 in the heavy chain and light chain variable regions of the antibody. In some embodiments, the dual inhibitory antibody specifically binds to two different proteins (e.g., KLK5 and KLK7) expressed from two different genes.

[0040] Effective dose: As used herein, “effective dose” refers to the amount of each activator (e.g., anti-KLK5 / KLK7+IL-4 bispecific antibody) required, either alone or in combination with one or more other activators, to confer a desired effect (e.g., a therapeutic effect on a subject). In some embodiments, the therapeutic effect is reduced KLK5 and / or KLK7 activity and / or reduced IL-4 activity and / or alleviated disease (e.g., Netherton syndrome, eosinophilic esophagitis, and atopic dermatitis) or associated symptoms, such as improved barrier function.

[0041] Framework: As used herein, the terms “framework” or “framework sequence” refer to the sequence remaining after removing the CDRs from the variable region. Since the precise definition of a CDR sequence can be determined by various systems, the meaning of the framework sequence is also subject to different interpretations accordingly. The six CDRs (CDR-L1, CDR-L2, and CDR-L3 of the light chain, and CDR-H1, CDR-H2, and CDR-H3 of the heavy chain) divide the framework regions of the light and heavy chains into four sub-regions (FR1, FR2, FR3, and FR4) on each chain, with CDR1 located between FR1 and FR2, CDR2 between FR2 and FR3, and CDR3 between FR3 and FR4. When a framework region is referred to by others without specifying a particular sub-region as FR1, FR2, FR3, or FR4, it represents a combination of FRs within the variable region of a single naturally occurring immunoglobulin chain. As used herein, FR represents one of four subregions, and multiple FRs represent two or more of the four subregions that constitute a framework region. Human heavy and light chain acceptor sequences are known in the art. In one embodiment, acceptor sequences known in the art may be used in the antibodies disclosed herein.

[0042] Human Antibodies: As used herein, the term “human antibody” is intended to include antibodies having variable and constant regions derived from human germline immunoglobulin sequences. The human antibodies of this disclosure may include amino acid residues not encoded by human germline immunoglobulin sequences (e.g., mutations introduced by random or site-directed mutagenesis in vitro, or by somatic mutation in vivo), for example, in the CDR, specifically CDR3. However, as used herein, the term “human antibody” is not intended to include antibodies in which a CDR sequence derived from the germline of another mammalian species, such as mouse, has been transplanted into a human framework sequence.

[0043] Humanized Antibodies: As used herein, the term “humanized antibody” refers to an antibody that contains heavy and light chain variable region sequences derived from a non-human species (e.g., mouse), but in which at least a portion of the VH and / or VL sequences has been modified to be more “human-like,” i.e., more similar to human germline variable sequences. One type of humanized antibody is a CDR-transplant antibody in which a human CDR sequence is introduced into non-human VH and VL sequences to replace the corresponding non-human CDR sequence. In one embodiment, a humanized antibody is provided. Such an antibody may be produced by obtaining a mouse monoclonal antibody using conventional hybridoma technology and then humanizing it using in vitro genetic engineering, such as that disclosed in PCT Publication WO2005 / 123126A2 by Kasaian et al.

[0044] A humanized antibody is a human immunoglobulin (recipient antibody) in which residues from the recipient's complementarity-determining region (CDR) are replaced with residues from the CDR of a non-human species (donor antibody), such as mouse, rat, or rabbit, having the desired specificity, affinity, and capabilities. In some embodiments, Fv framework region (FR) residues of the human immunoglobulin are replaced with corresponding non-human residues. Furthermore, the humanized antibody may include residues not found in the recipient antibody or the transferred CDR or framework sequence, but included to further improve and optimize the antibody's performance. Generally, a humanized antibody may include substantially all of at least one, typically two, variable domains, where all or substantially all of the CDR region corresponds to that of a non-human immunoglobulin, and all or substantially all of the FR region corresponds to that of the human immunoglobulin consensus sequence. The humanized antibody also includes, optimally, at least a portion of the immunoglobulin constant region or domain (Fc), typically at least a portion of the constant region or domain (Fc) of the human immunoglobulin. Antibodies may have modified Fc regions as described in WO99 / 58572. Other forms of humanized antibodies have one or more modified CDRs (1, 2, 3, 4, 5, 6) relative to the original antibody, also called one or more CDRs derived from one or more CDRs of the original antibody. Humanized antibodies may also undergo affinity maturation.

[0045] In some embodiments, humanization is achieved by transplanting a CDR (e.g., those shown in Table 1a or 1b) into a human variable domain (e.g., IGKV1-NL1*01 and IGHV1-3*01 human variable domains). In some embodiments, the antibodies of this disclosure are humanized variants comprising one or more amino acid substitutions (e.g., in the VH framework region) compared to any one of the VHs listed in Table 1a or 1b, and / or one or more amino acid substitutions (e.g., in the VL framework region) compared to any one of the VLs listed in Table 1a or 1b.

[0046] Isolated Antibodies: As used herein, “isolated antibodies” are intended to refer to antibodies that substantially contain no other antibodies with different antigen specificities. However, isolated antibodies may, in some embodiments, exhibit cross-reactivity to other antigens. Furthermore, isolated antibodies may substantially contain no other cellular material and / or chemical substances.

[0047] Kabat Numbering: As used herein, the terms “Kabat numbering,” “Kabat definition,” and “Kabat labeling” are used synonymously. These terms, as recognized in the art, refer to a system for numbering amino acid residues that are more variable than other amino acid residues (i.e., hypervariable) in the heavy chain variable region and light chain variable region of an antibody, or in their antigen-binding regions (Kabat et al. (1971) Ann. NY Acad, Sci. 190:382-391 and Kabat, EA, et al. (1991) Sequences of Proteins of Immunological Interest, Fifth Edition, USD Department of Health and Human Services, NIH Publication No. 91-3242). In the heavy chain variable region, the hypervariable region ranges from amino acid positions 31-35 in CDR1, from amino acid positions 50-65 in CDR2, and from amino acid positions 95-102 in CDR3. In the case of the heavy chain variable region, the hypervariable region is located at amino acid positions 24-34 in CDR1, amino acid positions 50-56 in CDR2, and amino acid positions 89-97 in CDR3.

[0048] Multispecific antigen-binding molecule: As used herein, the term “multispecific antigen-binding molecule” refers to a molecule containing two or more antigen-specific binding sites. In some embodiments, the multispecific antigen-binding molecule is a multispecific antibody (e.g., a bispecific antibody).

[0049] Multispecific antibody: As used herein, the term “multispecific antibody” refers to an antibody comprising at least two distinct antigen-specific binding sites, or, in combination, at least two (covalently or noncovalently) linked antibodies comprising at least two distinct antigen-specific binding sites. In some embodiments, a multispecific antibody is a bispecific antibody. Non-limiting examples of the format or structure of multispecific antibodies are provided in Sawant MS, et al., Toward Drug-Like Multispecific Antibodies by Design, Int J Mol Sci. 2020 Oct 12;21(20):7496; Klein C, et al., The use of CrossMAb technology for the generation of bi- and multispecific antibodies, MAbs 2016 Aug-Sep;8(6):1010-20; and Brinkmann U and Kontermann EE, The making of bispecific antibodies, MAbs. 2017 Feb / Mar;9(2):182-212, the entire contents of each of these are incorporated herein by reference.

[0050] Recombinant Antibodies: As used herein, the term “recombinant antibodies” means, for example, antibodies isolated from recombinant, combined human antibody libraries (Hoogenboom HR, (1997) TIB Tech. 15:62-70, Azzazy H., and Highsmith WE, (2002) Clin. Biochem. 35:425-445, Gavilondo JV, and Larrick JW (2002) BioTechniques 29:128-145, Hoogenboom H., and Chames P. (2000) Immunology Today 21:371-378), or antibodies isolated from animals transgenic to human immunoglobulin genes (e.g., mice) (e.g., Taylor, LD, et al. (1992) Nucl. Acids Res. 20:6287-6295, Kellermann SA., and Green LL (2002) Current Opinion). The disclosure is intended to include all antibodies prepared, expressed, produced, or isolated by recombinant means, such as antibodies expressed using a recombinant expression vector transfected into host cells (as described in detail herein), including antibodies prepared, expressed, produced, or isolated by any other means involving splicing of human immunoglobulin gene sequences against other DNA sequences (see in Biotechnology 13:593-597, Little M. et al (2000) Immunology Today 21:364-370), or by any other means involving splicing of human immunoglobulin gene sequences against other DNA sequences. In some embodiments, recombinant human antibodies are provided herein. In certain embodiments, such recombinant human antibodies have a variable region and a constant region derived from a human germline immunoglobulin sequence. However, in certain embodiments, such recombinant human antibodies are subjected to in vitro mutagenesis (or, if animal transgenicity for human Ig sequences is used, in vivo somatic mutagenesis), and therefore the amino acid sequences of the VH and VL regions of the recombinant antibody are derived from human germline VH and VL sequences, but may not be naturally present in vivo within the human antibody germline repertoire.One embodiment of the present disclosure provides, for example, a complete human antibody capable of binding to human KLK5 or KLK7, which can be generated using appropriate techniques, such as using a human Ig phage library, such as that disclosed in Jermutus et al., PCT Publication WO2005 / 007699A2.

[0051] Selective: As used herein, the terms “selective” or “selectively” refer to the ability of a molecule to produce an effect (e.g., inhibition, antagonism, agonization, etc.) on its target molecule compared to a reference molecule. For example, a molecule that selectively inhibits its target molecule means that this molecule can inhibit the target molecule to a degree that is distinguishable from the reference molecule in an inhibition assay or other inhibition situation. For example, with respect to an inhibitor, the term “selectively inhibits” refers to the ability of the inhibitor to inhibit the target molecule to a degree that is distinguishable from a reference molecule that does not substantially inhibit it in an inhibition assay, to a degree that enables selective inhibition of the target molecule, as described herein. After the reaction is complete, the signal produced by the inhibition of the target molecule can be measured. The maximum half-volume inhibitor concentrations for the target molecule and the reference molecule can be calculated.

[0052] Specific binding: As used herein, the term “specifically binding” refers to the ability of a molecule to bind to a binding partner with a degree of affinity or binding strength that enables the molecule to be used to distinguish the binding partner from a suitable control in a binding assay or other binding situation. With respect to an antibody, the term “specifically binding” refers to the ability of an antibody to bind to a specific antigen with a certain degree of affinity or binding strength compared to a suitable reference antigen(s), as described herein, enabling the antibody to be used to distinguish a particular antigen from other antigens. In some embodiments, the K of the antibody to bind to a target D However, at least about 10 -4 M, 10 -5 M, 10 -6 M, 10 -7 M, 10 -8M, 10 -9 M, 10 -10 M, 10 -11 M, 10 -12 M, 10 -13 When M is 10 or less, the antibody binds specifically to the target. In some embodiments, the antibody binds specifically to KLK5 or KLK7.

[0053] Subject: As used herein, the term "subject" refers to a mammal. In some embodiments, the subject is a non-human primate or a rodent. In some embodiments, the subject is a human. In some embodiments, the subject is a patient having or suspected of having a disease, e.g., a human patient.

[0054] Treatment: As used herein, the terms "treat" or "treatment" refer to the application or administration of a composition comprising one or more active agents (e.g., an anti-KLK5 / KLK7 + IL-4 bispecific antibody) to a subject having a target disease or disorder, a disease / disorder symptom, or a predisposition to a disease or disorder, for the purpose of curing, healing, alleviating, mitigating, modifying, correcting, ameliorating, improving, or affecting the disease, disease symptom, or predisposition. Alleviation of a target disease / disorder includes delaying or preventing the onset or progression of the disease or reducing the severity of the disease. It is understood that a reference to treating or treatment may also refer to an antibody comprising a KLK5 / KLK7 + IL-4 bispecific antibody for use in such methods.

[0055] II. Antibody (a) Dual inhibitory KLK5 and KLK7 antibodies In some embodiments, dual inhibitory antibodies targeting KLK5 and KLK7 (referred to as anti-KLK5 / KLK7 antibodies) are antibodies specific to both kallikrein-5 (KLK5) and KLK7 via a common specific antigen-binding site. Dual inhibitory KLK5 / KLK7 antibodies are described in PCT / US2024 / 019231, the entire contents of which are incorporated herein by reference. In some embodiments, antibodies are provided herein that conjugate with high specificity and affinity to KLK5 (e.g., human KLK5 or mouse KLK5) and KLK7 (e.g., human KLK7 or mouse KLK7) via a common antigen-binding site. In some embodiments, the anti-KLK5 / KLK7 antibodies described herein specifically conjugate to the KLK5 epitope that is or will be exposed to the antibody, and to the KLK7 epitope that is or will be exposed to the antibody. In some embodiments, the anti-KLK5 / KLK7 antibody described herein binds to the active sites of KLK5 and KLK7. In some embodiments, the anti-KLK5 / KLK7 antibody provided herein specifically binds to KLK5 and KLK7 from humans, non-human primates, mice, rats, etc. In some embodiments, the anti-KLK5 / KLK7 antibody provided herein specifically binds to human KLK5. In some embodiments, the anti-KLK5 / LKL7 antibody provided herein specifically binds to mouse KLK5. In some embodiments, the anti-KLK5 / KLK7 antibody provided herein specifically binds to human KLK7. In some embodiments, the anti-KLK5 / LKL7 antibody provided herein specifically binds to mouse KLK7. In some embodiments, a multispecific antibody is provided herein, comprising an arm having an antigen-specific binding site for a dual inhibitory antibody targeting KLK5 and KLK7, and an arm having an antigen-specific binding site that specifically binds to IL-4. For example, in some embodiments, a bispecific antibody is provided herein that includes an arm having an antigen-specific binding site for a dual inhibitory antibody targeting KLK5 and KLK, and an arm having an antigen-specific binding site that specifically binds to IL-4 or IL-4R.

[0056] In some embodiments, the anti-KLK5 / KLK7 antibody is not a bispecific antibody or bispecific antigen-binding molecule in which KLK5 binding is conferred by one binding site within the antibody and KLK7 binding is conferred by a different binding site within the antibody.

[0057] In some embodiments, the anti-KLK5 / KLK7 antibodies described herein may be characterized by reference to certain functional properties. In some embodiments, the anti-KLK5 / KLK7 antibodies described herein specifically bind to KLK5 and KLK7. In some embodiments, the anti-KLK5 / KLK7 antibodies specifically bind to the active forms of KLK5 and KLK7. In some embodiments, the anti-KLK5 / KLK7 antibodies do not bind to the inactive forms (proforms) of KLK5 and KLK7. In some embodiments, the antibody specifically binds to the active forms of KLK5 and KLK7, but does not specifically bind to the inactive forms of KLK5 or KLK7. In some embodiments, the antibody detectably binds to the active forms of KLK5 and KLK7, but does not detectably bind to the inactive forms of KLK5 or KLK7 under the same or equivalent conditions. In some embodiments, anti-KLK5 / KLK7 antibodies inhibit KLK5 and KLK7 protease activity. In some embodiments, anti-KLK5 / KLK7 antibodies are not cleaved by KLK5 or KLK7 when bound to them. In some embodiments, anti-KLK5 / KLK7 antibodies compete with SPINK5 and / or leupeptin for binding to the active sites of KLK5 and KLK7. In some embodiments, anti-KLK5 / KLK7 antibodies reduce keratosis and desquamation. In some embodiments, anti-KLK5 / KLK7 antibodies reduce stratum corneum thickness. In some embodiments, anti-KLK5 / KLK7 antibodies reduce inflammation and epidermal effects.

[0058] Kallikrein-5, also known as stratum corneum trypsinase (SCTE), is a serine protease expressed in the epidermis, encoded by the KLK5 gene. The KLK5 gene is one of 15 kallikrein subfamily members located in a cluster on the chromosome. Its expression is upregulated by estrogen and progestin. KLK5 is expressed in the stratum granulosum and stratum corneum. In some embodiments, KLK5 controls epidermal desquamation. In some embodiments, KLK5 controls epidermal desquamation in conjunction with other members of the kallikrein family proteases (e.g., KLK7 and / or KLK14). In some embodiments, KLK5 degrades proteins that make up the epidermis (e.g., stratum corneum, stratum clearusa, stratum granulosum, stratum spinosum, or stratum basale). In some embodiments, KLK5 degrades proteins that form the stratum corneum and / or granular layer (e.g., cornodesmosine (CDSN), desmoglein 1 (DSG1), and desmocolin 1 (DSC1)). In the epidermis (e.g., granular layer and stratum corneum), KLK5 is expressed as pro-KLK5, an inactive form (sometimes called proform or pro-form), which can self-activate itself. Once activated, KLK5 can convert both pro-KLK7 and pro-KLK14 to their active forms via proteolytic cleavage. Subsequently, active KLK14 can activate newly generated pro-KLK5, thus forming a positive feedback loop (see, for example, Nauroy et al., Kallikreins: Essential epidermal messengers for regulation of the skin microenvironment during homeostasis, repair and disease, Matrix Biol Plus. 2019;6-7:100019). KLK7 and KLK14 also degrade proteins that form the stratum corneum and / or granular layer (e.g., cornodesmosine (CDSN), desmoglein 1 (DSG1), and desmocolin 1 (DSC1)).Structural proteins such as CDSN, DSG1, and DSC1 are adhesion proteins in the extracellular portion of corneodesmosomes, which are junctional structures that mediate the adhesion of keratinocytes. Degradation of these proteins on the epidermal surface leads to desquamation, which can result in skin barrier defects (e.g., stratum corneum exfoliation, decreased permeability barrier, allergies, and inflammation). KLK5 and KLK7 are involved in this process (see, for example, Caubet et al., Degradation of Corneodesmosome Proteins by Two Serine Proteases of the Kallikrein Family, SCTE / KLK5 / hK5 and SCCE / KLK7 / hK7, Journal of Investigative Dermatology, Volume 122, Issue 5, May 2004, Pages 1235-1244). Inhibition of KLK5 and / or KLK7 promotes improved skin barrier integrity and reduced inflammation (e.g., Chavarria-Smith et al., Dual antibody inhibition of KLK5 and KLK7 for Netherton syndrome and atopic dermatitis, SCIENCE TRANSLATIONAL MEDICINE, 14 Dec 2022, Vol 14, Issue 675).

[0059] Kallikrein-7 is a serine protease encoded by the KLK7 gene in humans. KLK7 is characterized as a stratum corneum chymotriptic enzyme (SCCE). It is the seventh member of the human kallikrein family, which includes 15 homologous serine proteases located on chromosome 19. KLK7 is secreted as an inactive zymogen (e.g., in the granular layer of the epidermis) and requires proteolytic cleavage to be activated. In some embodiments, KLK5 or matryptase activates KLK7. Once activated, KLK7 can cleave proteins that form the stratum corneum and / or granular layer (e.g., corneodesmosine (CDSN), desmoglein 1 (DSG1), and desmocolin 1 (DSC1)) (see, for example, Caubet et al. (May 2004). Degradation of corneodesmosome proteins by two serine proteases of the kallikrein family, SCTE / KLK5 / hK5 and SCCE / KLK7 / hK7. The Journal of Investigative Dermatology. 122(5):1235-1244). These proteins constitute the extracellular component of corneodesmosomes, which are intercellular adhesion structures that connect intermediate filaments of adjacent cells in the stratum corneum. In some embodiments, proteolysis of keratinodesmosomes leads to epidermal desquamation (i.e., the shedding of keratinocytes from the outer layer of the epidermis). In some embodiments, the combined roles of KLK5 and KLK7 suggest that the KLK cutaneous cascade is responsible for regulating desquamation. KLK7 is a chymotrypsin-like serine protease that cleaves proteins at tyrosine, phenylalanine, or leucine residues. In some embodiments, dysregulation of KLK7 is linked to several skin disorders, including atopic dermatitis, psoriasis, and Netherton syndrome. These diseases are characterized by excessively dry, scaly, and inflamed skin due to a breakdown of skin homeostasis and normal barrier function.

[0060] In some embodiments, the anti-KLK5 / KLK7 antibodies described herein specifically bind to epitopes on human KLK5. Exemplary amino acid sequences of human KLK5 are described in NCBI accession numbers NP_001070959.1, NP_001070960.1, or NP_036559.1, and UniProt accession numbers: Q8IU55, Q6S9W8, M0QXX2, Q9P0G3, A0A2I2MP48, or A0A2I2MP49, the full sequences of which are incorporated herein by reference.

[0061] In some embodiments, the anti-KLK5 / KLK7 antibodies described herein specifically bind to epitopes on mouse KLK5. Exemplary amino acid sequences of mouse KLK5 are described in NCBI accession numbers NP_081082.1, XP_006541213.1, XP_006541214.1, XP_006541215.1, XP_036009294.1, or XP_036009295.1, and UniProt accession number P15945, or Q9D140, the full sequences of which are incorporated herein by reference.

[0062] In some embodiments, the anti-KLK5 / KLK7 antibodies described herein specifically bind to an epitope on human KLK7 via the same antigen-binding site that binds to KLK5 (e.g., human KLK5 or mouse KLK5). Exemplary amino acid sequences of human KLK7 are described in NCBI accession numbers NP_001193982.1, NP_001230055.1, NP_005037.1, NP_644806.1, and UniProt accession numbers: M0QYU8, Q6DTY1, X2J289, X2J4X7, A0A024R4H6, P49862, A0A2H4GDB2, and A0A2H4GDB6, the full sequences of which are incorporated herein by reference.

[0063] In some embodiments, the anti-KLK5 / KLK7 antibodies described herein specifically bind to an epitope on mouse KLK7 via the same antigen-binding site that binds to KLK5 (e.g., human KLK5 or mouse KLK5). Exemplary amino acid sequences of mouse KLK7 are described in NCBI accession number NP_036002.1 and UniProt accession number Q91VE3, and these full sequences are incorporated herein by reference.

[0064] In some embodiments, the anti-KLK5 / KLK7 antibody described herein specifically binds to an epitope on KLK5 (e.g., the active site of the enzyme, also known as the catalytic domain / pocket of human KLK5 or mouse KLK5) and an epitope on KLK7 (e.g., the catalytic domain / pocket of human KLK7 or mouse KLK7). In some embodiments, the anti-KLK5 / KLK7 antibody described herein prevents KLK5 (e.g., human or mouse KLK5) and KLK7 (e.g., human or mouse KLK7) from cleaving their substrates. In some embodiments, the anti-KLK5 antibody described herein binds to fragments of KLK5 (e.g., human or mouse KLK5) and fragments of KLK7 (e.g., human or mouse KLK7). Fragments of KLK5 and / or KLK7 (e.g., human or mouse) may be approximately 5 to 425 amino acids, 10 to 400 amino acids, 50 to 350 amino acids, 100 to 300 amino acids, 150 to 250 amino acids, 200 to 300 amino acids, 75 to 150 amino acids, 25 to 100 amino acids, or 10 to 30 amino acids in length. Without intending to be bound by any particular theory, in some embodiments, the heavy chain (HC) complementarity determining region 3 (CDR3) of any one of the anti-KLK5 / KLK7 antibodies described herein inhibits KLK5 (e.g., human or mouse KLK5) and KLK7 (e.g., human or mouse KLK7) by binding to the catalytic domain / pocket of KLK5.

[0065] In some embodiments, the anti-KLK5 / KLK7 antibodies described herein inhibit KLK5 protease activity, KLK7 protease activity, or protease activity of both KLK5 and KLK7. In some embodiments, the anti-KLK5 / KLK7 antibodies inhibit KLK5 cleavage of BOC-Val-Pro-Arg-AMC with an IC50 of less than 30 nM, less than 25 nM, less than 20 nM, less than 15 nM, less than 10 nM, less than 5 nM, less than 3 nM, less than 2.5 nM, less than 2 nM, or less than 1.5 nM, less than 1 nM, less than 0.5 nM, less than 0.3 nM, less than 0.25 nM, less than 0.2 nM, or less than 0.1 nM. In some embodiments, the anti-KLK5 / KLK7 antibody is expressed in the following concentrations: 0.1nM-30nM, 0.1nM-20nM, 0.1nM-10nM, 0.1nM-5nM, 0.1nM-2.5nM, 0.1nM-2nM, 0.1nM-1nM, 0.1nM-0.5nM, 0.1nM-0.25nM, 0.1nM-50nM, 0.1nM-40nM, 0.1nM-30nM, 0.1nM-20nM, 0.1nM-10nM. M, 0.1nM~5nM, 0.1nM~2.5nM, 0.1nM~2nM, 0.1nM~1nM, 0.1nM~0.9nM, 0.1nM~0.8nM, 0.1nM~0.7nM, 0.1nM~0. 6nM, 0.1nM~0.5nM, 0.1nM~0.4nM, 0.1nM~0.3nM, 0.1nM~0.25nM, 0.1nM~0.2nM, 0.1nM~0.15nM, 0.15nM~0.2n M, 0.15nM~0.25nM, 0.15nM~0.3nM, 0.15nM~0.4nM, 0.15nM~0.5nM, 0.15nM~1nM, 0.2nM~30nM, 0.2nM~20nM, 0.2nM~10nM, 0.2nM~5nM, 0.2nM~2.5nM, 0.2nM~2nM, 0.2nM~1nM, 0.2nM~0.5nM, 0.2nM~0.2nM, 0.2nM~50nM, 0 .2nM~40nM, 0.2nM~30nM, 0.2nM~20nM, 0.2nM~10nM, 0.2nM~5nM, 0.2nM~2.5nM, 0.2nM~2nM, 0.2nM~1nM, 0.2 nM~0.9nM, 0.2nM~0.8nM, 0.2nM~0.7nM, 0.2nM~0.6nM, 0.2nM~0.5nM, 0.2nM~0.4nM, 0.2nM~0.3nM, 0.2nM~0.25nM, 1nM~30nM, 1nM~20nM, 1nM~10nM, 1nM~5nM, 1nM~2.5nM, 1nM~2nM, 1nM~3nM, 1nM~5.5nM, 1.5nM~2nM, 1.5nM~3nM, 1.5nM~5.5nM, 2nM~5nM, 2nM~4nM, 2nM~5.5nM, 3nM~5.5nM, 4nM~5.5nM, 3nM~30nM, 3nM~20nM, 3 nM~10nM, 3nM~5nM, 3nM~2.5nM, 3nM~4nM, 3nM~5.5nM, 5nM~30nM, 5nM~20nM, 5nM~10nM, 5nM~9nM, 5nM~8nM , 5nM~7nM, 5nM~6nM, 5nM~5.5nM, 10nM~30nM, 10nM~25nM, 10nM~20nM, 10nM~18nM, 10nM~15nM, 10nM~12nM, 12nM~20nM, 12nM~25nM, 12nM~16nM, 12nM~18nM, 12nM~20nM, 12nM~24nM, 12nM~28nM, 12nM~30nM, 15nM~3 0nM, 15nM~25nM, 15nM~20nM, 15nM~18nM, 18nM~30nM, 18nM~25nM, 18nM~20nM, 20nM~30nM, 20nM~25nM, 20n It inhibits the cleavage of KLK5 (e.g., human KLK5 or mouse KLK5) in BOC-Val-Pro-Arg-AMC with an IC50 in the range of M~22nM, 20nM~24nM, 20nM~26nM, 22nM~30nM, 22nM~25nM, 22nM~28nM, 24nM~30nM, 24nM~25nM, 24nM~26nM, or 24nM~28nM. In some embodiments, the anti-KLK5 / KLK7 antibody inhibits KLK7 cleavage of KHLF-AMC with an IC50 of less than 6 nM, less than 5 nM, less than 4 nM, less than 3 nM, less than 2.5 nM, less than 2 nM, or less than 1.5 nM, less than 1 nM, less than 0.5 nM, less than 0.4 nM, less than 0.3 nM, less than 0.2 nM, less than 0.16 nM, less than 0.1 nM, or less than 0.05 nM. In some embodiments, the anti-KLK5 / KLK7 antibody is expressed in concentrations of 0.1nM to 30nM, 0.1nM to 20nM, 0.1nM to 10nM, 0.1nM to 5nM, 0.1nM to 2.5nM, 0.1nM to 2nM, 0.1nM to 1nM, 0.1nM to 0.5nM, 0.1nM to 0.25nM, 0.1nM to 50nM, and 0.1nM~40nM、0.1nM~30nM、0.1nM~20nM、0.1nM~10nM、0.1nM~5nM、0.1nM~2.5nM、0.1nM~2nM、0.1nM~1nM、0.1nM~0.9nM、0.1nM~0.8nM、0.1nM~0.7nM、0.1nM~0.6nM、0.1nM~0.5nM、0.1nM~0.4nM、0.1nM~0.3nM、0.1nM~0.25nM、0.1nM~0.2nM、0.1nM~0.15nM、0.15nM~0.2nM、0.15nM~0.25nM、0.15nM~0.3nM、0.15nM~0.4nM、0.15nM~0.5nM、0.15nM~1nM、0.2nM~30nM、0.2nM~20nM、0.2nM~10nM、0.2nM~5nM、0.2nM~2.5nM、0.2nM~2nM、0.2nM~1nM、0.2nM~0.5nM、0.2nM~0.2nM、0.2nM~50nM、0.2nM~40nM、0.2nM~30nM、0.2nM~20nM、0.2nM~10nM、0.2nM~5nM、0.2nM~2.5nM、0.2nM~2nM、0.2nM~1nM、0.2nM~0.9nM、0.2nM~0.8nM、0.2nM~0.7nM、0.2nM~0.6nM、0.2nM~0.5nM、0.2nM~0.4nM、0.2nM~0.3nM、0.2nM~0.25nM、1nM~30nM、1nM~20nM、1nM~10nM、1nM~5nM、1nM~2.5nM、1nM~2nM、1nM~3nM、1nM~5.5nM、1.5nM~2nM、1.5nM~3nM、1.5nM~5.5nM、2nM~5nM、2nM~4nM、2nM~5.5nM、3nM~5.5nM、4nM~5.5nM、3nM~30nM、3nM~20nM、3nM~10nM、3nM~5nM、3nM~2.5nM、3nM~4nM、3nM~5.5nM、5nM~30nM、5nM~20nM、5nM~10nM、5nM~9nM、5nM~8nM、5nM~7nM、5nM~6nM、5nM~5.5nM, 10nM~30nM, 10nM~25nM, 10nM~20nM, 10nM~18nM, 10nM~15nM, 10nM~12nM, 12nM~20nM, 12nM~25nM, 12nM~16nM, 12nM~18nM, 12nM~20nM, 12nM~24nM, 12nM~28nM, 12nM~30nM, 15nM~30nM, 15nM~25nM, 15nM~20nM, 15nM~18nM, 18nM IC50 values ​​in the ranges of ~30nM, 18nM~25nM, 18nM~20nM, 20nM~30nM, 20nM~25nM, 20nM~22nM, 20nM~24nM, 20nM~26nM, 20nM~28nM, 22nM~30nM, 22nM~25nM, 22nM~28nM, 24nM~30nM, 24nM~25nM, 24nM~26nM, or 24nM~28nM inhibit KLK7 cleavage of KHLF-AMC.

[0066] In some embodiments, the anti-KLK5 / KLK7 antibody described herein specifically binds to KLK5, KLK7, or the active forms of KLK5 and KLK7. In some embodiments, the anti-KLK5 / KLK7 antibody described herein does not bind to KLK5, KLK7, or the inactive forms of KLK5 and KLK7. In some embodiments, the anti-KLK5 / KLK7 antibody described herein specifically binds to the active site of KLK5, KLK7, or KLK5 and KLK7. In some embodiments, the anti-KLK5 / KLK7 antibody described herein specifically binds to the active form of KLK5, the active form of KLK7, or the active forms of KLK5 and KLK7, but does not specifically bind to the inactive form of KLK5, the inactive form of KLK7, or the inactive forms of KLK5 and KLK7. In some embodiments, the anti-KLK5 / KLK7 antibodies described herein are detectably bound to the active form of KLK5, the active form of KLK7, or the active forms of both KLK5 and KLK7, but are not detectably bound to the inactive form of KLK5, the inactive form of KLK7, or the inactive forms of both KLK5 and KLK7 under the same or equivalent conditions. The active site of KLK5 and / or KLK7 is the site on which the KLK5 and / or KLK7 substrate molecule binds and is cleaved. The active site may also be known as the catalytic domain or catalytic triresidue. In some embodiments, the active site of KLK5 or KLK7 (i.e., the catalytic domain or catalytic triresidue) consists of the amino acids Ser195, His57, and Asp102 of KLK5 or KLK7 (see, for example, Goettig et al., Natural and synthetic inhibitors of kallikrein-related peptidases (KLKs), Biochimie. 2010 Nov;92(11):1546-1567).

[0067] In some embodiments, the antibodies described herein are optimized versions (e.g., affinity-mature) of the parent antibody. In some embodiments, the antibodies described herein are at least about 10 -4 M, at least about 10 -5 M, at least about 10 -6 M, at least about 10-7 M, at least about 10 -8 M, at least about 10 -9 M, at least about 10 -10 M, at least about 10 -11 M, at least about 10 -12 M, at least about 10 -13 M, or lower binding affinity (e.g., K D The antibodies specifically bind to KLK5 (e.g., human or mouse KLK5) and KLK7 (e.g., human or mouse KLK7) as shown by [representation]. In some embodiments, the antibodies described herein bind to 1 × 10⁻⁶ antibodies. -10 M~5×10 -9 M, 1×10 -10 M~1×10 -9 M, 5×10 -10 ~1 × 10 -9 M, 5×10 -11 ~1 × 10 -10 M, 1×10 -11 ~5×10 -10 M, or 5×10 -13 ~1 × 10 -12 Binding affinity of M (e.g., K DThe antibodies of this disclosure specifically bind to KLK5 (e.g., human or mouse KLK5) and KLK7 (e.g., human or mouse KLK7) with affinities of 1 pM to 500 nM, e.g., 50 pM to 100 nM, 500 pM to 50 nM, 1 pM to 100 pM, 10 pM to 100 pM, 50 pM to 100 pM, 100 pM to 500 pM, 500 pM to 1 nM, 1 nM to 5 nM, 1 nM to 10 nM, 5 nM to 25 nM, 10 nM to 50 nM, 50 nM to 100 nM, and 100 nM to 500 nM. This disclosure also includes antibodies that compete with any of the antibodies described herein for binding to KLK5 protein (e.g., human or mouse KLK5) and KLK7 protein (e.g., human or mouse KLK7) and have affinities of 100 nM or less (e.g., 80 nM or less, 50 nM or less, 20 nM or less, 10 nM or less, 1 nM or less, 500 pM or less, 50 pM or less, or 5 pM or less). The affinity and binding reaction rate of the antibodies can be tested using any suitable method, including but not limited to biosensor technology (e.g., OCTET or BIACORE). In some embodiments, the antibodies described herein have K in the sub-nanomole range. D It connects to KLK5 and KLK7.

[0068] Binding affinity (or binding specificity) can be determined by a variety of methods, including equilibrium dialysis, equilibrium binding, gel filtration, ELISA, surface plasmon resonance (SPR), fluorescence-activated cell sorting (FACS), or spectroscopy (e.g., using fluorescence assays). Exemplary conditions for evaluating binding affinity are HBS-P buffer (10 mM HEPES pH 7.4, 150 mM NaCl, 0.005% (v / v) surfactant P2O) and PBS buffer (10 mM PO4-3, 137 mM NaCl, and 2.7 mM KCl). Using these techniques, the concentration of the binding protein can be measured as a function of the target protein concentration. Generally, the concentration of the binding protein ([[binding]]) is given by the following formula: The binding rate is related to the concentration of the free target protein (free protein) because [[binding]] = [[free]] / (Kd + [[free]]).

[0069] K A While it is not always necessary to make an accurate determination, obtaining a quantitative measure of affinity determined using methods such as ELISA or FACS analysis is useful if it is K A Since it is proportional to the activity, it may be sufficient, and therefore can be used for comparisons to determine whether a higher affinity is, for example, twice as high, to obtain a qualitative measurement of affinity, or to infer affinity by activity in a functional assay (e.g., an in vitro or in vivo assay).

[0070] Exemplary anti-KLK5 / KLK7 antibody sequences (e.g., heavy chain (HC) sequence and light chain (LC) sequence, heavy chain variable domain (VH) and light chain variable domain (VL), and CDR sequence) are provided in Tables 1a and 1b. [Table 1a-1] [Table 1a-2]

[0071] In some embodiments, certain amino acid positions in the antibodies described herein (e.g., amino acids within the VH / VL region and / or CDR region) are substitutable, and this substitution results in an antibody having substantially similar binding and biological activity (e.g., substantially similar binding affinity, binding specificity, protease inhibitory activity, anti-inflammatory activity, or a combination thereof) to the reference antibody. To identify substitutable positions in an antibody, the amino acid sequence of that antibody is compared to the sequences of other antibodies belonging to the same group as that antibody. If the identity of that amino acid varies among different related antibodies of a certain group at any particular position, that position is a substitutable position in the antibody. In other words, a substitutable position is a position where the identity of the amino acid varies among related antibodies. Positions containing constant amino acids are not substitutable positions.

[0072] In some embodiments, the above method may be used to provide a consensus antibody sequence. In such a consensus sequence, non-substitutable positions are indicated by the amino acids present at those positions, and substitutable positions are indicated by "X".

[0073] Depending on how the antibody is adopted, X may be a) any amino acid, b) any amino acid present at that position in any of the related antibodies within that group, or a conservatively substituted variant thereof, or c) any amino acid present at that position in any of the related antibodies within that group. Any antibody with a sequence encompassed by consensus should bind to the same antigen as any of the related antibodies.

[0074] In some embodiments, the methods described above may be employed in a method for designing and producing variants of a parent antibody that at least maintain (e.g., maintain or increase) the antigen-binding activity of the parent antibody. Antibodies containing substitutions at substitutable positions can be produced and tested, and therefore substitutions at those positions should not significantly reduce the antibody's binding activity. Generally, antibody variants of a parent antibody have an antigen-binding affinity of at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 100% (e.g., at least 150%, at least 200%, at least 500%, at least 1000%, and typically up to at least 10,000%) of the parent antibody's binding affinity to a particular antigen.

[0075] In some embodiments, the substitutable positions of the parent antibody may be substituted by: a) any of 20 naturally occurring amino acids to produce a random substitution; b) an amino acid with biochemical properties similar to an amino acid already present at the substitutable position to produce a conservative substitution; c) an amino acid present at the same position in the related antibody to produce a designated substitution; or d) an amino acid present at the same position in a similar human antibody to produce a humanization substitution. Substitutions may occur in any portion of the antibody variable region, including any framework region or CDR. In certain embodiments, a single substitutable amino acid may be substituted. However, in other embodiments, multiple substitutable amino acids (e.g., up to about 5 or 10 or more) may be substituted. In certain embodiments, the type of substitution that can be performed at each substitutable position may be indicated by the type of amino acid present at that position in the related antibody. For example, if unrelated amino acids (e.g., Ala, Gly, Cys, Glu, and Thr) are present at certain positions in the group of related antibodies, any amino acid can be substituted at that position without significantly reducing the antibody's binding activity. Exemplary amino acid substitutions of anti-KLK5 / KLK7 antibodies described herein are shown in Table 1b: [Table 1b]

[0076] In some embodiments, the antibodies of this disclosure include HC CDR1 containing the amino acid sequence GSISSX1DYYWX2 (SEQ ID NO: 28) (where X1 is S, D, or L, and X2 is G or V), HC CDR2 containing the amino acid sequence SIX3YX4X5X6TYYX7PSLKS (SEQ ID NO: 29) (where X3 is Y or D, X4 is S, F, or Y, X5 is G or A, X6 is S or D, or X7 is N or S), HC CDR3 containing the amino acid sequence ARGRPLGYGAX8HX9YYGMDV (SEQ ID NO: 30) (where X8 is R or K, or X9 is Y or D), LC CDR1 containing the amino acid sequence of SEQ ID NO: 4, LC CDR2 containing the amino acid sequence of SEQ ID NO: 5, and / or QQSPX 10 FPPLT(Sequence ID 31)(X 10 It contains LC CDR3 with an amino acid sequence that is P or Y.

[0077] In some embodiments, the antibody of this disclosure comprises one or more HC CDR (e.g., HC CDR1, HC CDR2, or HC CDR3) amino acid sequences from any one of the anti-KLK5 / KLK7 antibodies selected from Tables 1a and 1b. In some embodiments, the antibody of this disclosure comprises an HC CDR3 amino acid sequence from any one of the anti-KLK5 / KLK7 antibodies selected from Tables 1a and 1b. In some embodiments, the antibody of this disclosure comprises HC CDR1, HC CDR2, and HC CDR3 provided for any one of the (elected) antibodies selected from Tables 1a and 1b. In some embodiments, the antibody of this disclosure comprises an LC CDR3 amino acid sequence from any one of the anti-KLK5 / KLK7 antibodies selected from Tables 1a and 1b. In some embodiments, the antibody of the Disclosure comprises one or more LC CDR (e.g., LC CDR1, LC CDR2, or LC CDR3) amino acid sequences from any one of the anti-KLK5 / KLK7 antibodies selected from Tables 1a and 1b. In some embodiments, the antibody of the Disclosure comprises LC CDR1, LC CDR2, and LC CDR3 provided for any one of the anti-KLK5 antibodies selected from Tables 1a and 1b.

[0078] In some embodiments, the antibodies of this disclosure include HC CDR1, HC CDR2, HC CDR3, LC CDR1, LC CDR2, and LC CDR3 provided for any one of the anti-KLK5 / KLK7 antibodies selected from Tables 1a and 1b. In some embodiments, the heavy chain CDR3 domain and / or light chain CDR3 domain of the antibody may play a particularly important role in the antibody's binding specificity / affinity to the antigen. Therefore, the antibodies of this disclosure may include at least the heavy chain CDR3 and / or light chain CDR3 of any one of the anti-KLK5 / KLK7 antibodies selected from Tables 1a and 1b.

[0079] Any variant of the exemplary anti-KLK5 / KLK7 antibodies disclosed herein is also within the scope of this disclosure. The variant may contain one or more amino acid residue mutations in the VH and / or VL, or in one or more of the HC CDRs and / or one or more of the LC CDRs, compared to the reference antibody, but retain substantially similar binding activity and biological activity (e.g., substantially similar binding affinity, binding specificity, protease inhibitory activity, anti-inflammatory activity, or a combination thereof) to the reference antibody.

[0080] In some embodiments, the antibodies of the present disclosure have one or more CDR (e.g., HC CDR or LC CDR) sequences substantially similar to any of HC CDR1, HC CDR2, HC CDR3, LC CDR1, LC CDR2, and / or LC CDR3 from one of the anti-KLK5 / KLK7 antibodies selected from Tables 1a and 1b. In some embodiments, the positions of one or more CDRs along the VH (e.g., HC CDR1, HC CDR2, or HC CDR3) and / or VL (e.g., LC CDR1, LC CDR2, or LC CDR3) regions of the antibodies described herein can be mutated by only 1, 2, 3, 4, 5, or 6 amino acid positions, as long as specific binding to KLK5 (e.g., human or mouse KLK5) and KLK7 (e.g., human or mouse KLK7) is maintained (e.g., substantially maintained, for example, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95% of the binding of the original antibody from which it is derived). For example, in some embodiments, the position defining the CDR of any antibody described herein can be mutated by shifting the N-terminal and / or C-terminal boundary of the CDR by 1, 2, 3, 4, 5, or 6 amino acids relative to the CDR position of any one of the antibodies described herein, insofar as specific binding of KLK5 (e.g., human or mouse KLK5) and KLK7 (e.g., human or mouse KLK7) is maintained (e.g., substantially maintained, for example, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95% of the binding of the original antibody from which it originates).In another embodiment, the length of one or more CDRs along the VH (e.g., HC CDR1, HC CDR2, or HC CDR3) and / or VL (e.g., LC CDR1, LC CDR2, or LC CDR3) regions of the antibodies described herein may be mutated (e.g., shortened or lengthened) by only 1, 2, 3, 4, 5, or more amino acids, as long as immunospecific binding to KLK5 (e.g., human or mouse KLK5) and KLK7 (e.g., human or mouse KLK7) is maintained (e.g., substantially maintained, for example, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95% of the binding of the original antibody from which it is derived).

[0081] Therefore, in some embodiments, the HC CDR1, HC CDR2, HC CDR3, LC CDR1, LC CDR2, and / or LC CDR3 described herein may be 1, 2, 3, 4, 5 amino acids, or more shorter than one or more of the CDRs described herein (e.g., CDRS from any of the anti-KLK5 / KLK7 antibodies selected from Tables 1a and 1b), as long as specific binding to KLK5 (e.g., human or mouse KLK5) and KLK7 (e.g., human or mouse KLK7) is maintained (e.g., substantially maintained by, for example, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95% compared to the binding of the original antibody from which it is derived). In some embodiments, the HC CDR1, HC CDR2, HC CDR3, LC CDR1, LC CDR2, and / or LC CDR3 described herein may be 1, 2, 3, 4, 5 amino acids, or more longer than one or more of the CDRs described herein (e.g., CDRs from any of the anti-KLK5 / KLK7 antibodies selected from Tables 1a and 1b), provided that specific binding to KLK5 (e.g., human or mouse KLK5) and KLK7 (e.g., human or mouse KLK7) is maintained (e.g., substantially maintained by at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95% compared to the binding of the original antibody from which it is derived). In some embodiments, the amino portions of HC CDR1, HC CDR2, HC CDR3, LC CDR1, LC CDR2, and / or LC CDR3 described herein can be extended by 1, 2, 3, 4, 5 amino acids or more compared to one or more of the CDRs described herein (e.g., CDRs from any of the anti-KLK5 / KLK7 antibodies selected from Tables 1a and 1b), provided that specific binding to KLK5 (e.g., human or mouse KLK5) and KLK7 (e.g., human or mouse KLK7) is maintained (e.g., substantially maintained by at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95% compared to the binding of the original antibody from which it is derived).In some embodiments, the carboxyl portions of HC CDR1, HC CDR2, HC CDR3, LC CDR1, LC CDR2, and / or LC CDR3 described herein can be extended by 1, 2, 3, 4, 5 amino acids or more compared to one or more of the CDRs described herein (e.g., CDRs from any of the anti-KLK5 / KLK7 antibodies selected from Tables 1a and 1b), provided that specific binding to KLK5 (e.g., human or mouse KLK5) and KLK7 (e.g., human or mouse KLK7) is maintained (e.g., substantially maintained by at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95% compared to the binding of the original antibody from which it is derived). In some embodiments, the amino portion of HC CDR1, HC CDR2, HC CDR3, LC CDR1, LC CDR2, and / or LC CDR3 described herein may be shortened by 1, 2, 3, 4, 5 amino acids or more compared to one or more of the CDRs described herein (e.g., CDRs from any of the anti-KLK5 / KLK7 antibodies selected from Tables 1a and 1b), as long as specific binding to KLK5 (e.g., human or mouse KLK5) and KLK7 (e.g., human or mouse KLK7) is maintained (e.g., substantially maintained by at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95% compared to the binding of the original antibody from which it is derived).In some embodiments, the carboxyl portion of HC CDR1, HC CDR2, HC CDR3, LC CDR1, LC CDR2, and / or LC CDR3 described herein can be shortened by 1, 2, 3, 4, 5 amino acids or more compared to one or more of the CDRs described herein (e.g., CDRs from any of the anti-KLK5 / KLK7 antibodies selected from Tables 1a and 1b), as long as specific binding to KLK5 (e.g., human or mouse KLK5) and KLK7 (e.g., human or mouse KLK7) is maintained (e.g., substantially maintained by at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95% compared to the binding of the original antibody from which it is derived). Any method can be used to confirm whether specific binding to KLK5 (e.g., human or mouse KLK5) and KLK7 (e.g., human or mouse KLK7) is maintained, for example, by using the binding assays and conditions described in the Art.

[0082] In some examples, the antibodies of this disclosure have one or more CDR (e.g., HC CDR or LC CDR) sequences substantially similar to any one of the anti-KLK5 / KLK7 antibodies selected from Tables 1a and 1b. For example, the antibodies described herein may contain one or more CDR sequences from any of the anti-KLK5 / KLK7 antibodies selected from Tables 1a and 1b, containing up to 5, 4, 3, 2, or 1 amino acid residue mutations compared to the corresponding CDR region of any one of the CDRs provided herein (e.g., CDRs from any of the anti-KLK5 / KLK7 antibodies selected from Tables 1a and 1b), insofar as specific binding to KLK5 (e.g., human or mouse KLK5) and KLK7 (e.g., human or mouse KLK7) is maintained (e.g., substantially maintained by at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95% compared to the binding of the original antibody from which it is derived). In some embodiments, any amino acid mutation in any of the CDRs provided herein may be a conservative mutation. A conservative mutation can be introduced into a CDR at a position where the residue is unlikely to be involved in interaction with KLK5 (e.g., human or mouse KLK5) and / or KLK7 (e.g., human or mouse KLK7) when determined, for example, based on the crystal structure. Some aspects of this disclosure provide antibodies comprising one or more heavy chain variable (VH) domains and / or light chain variable (VL) domains provided herein. In some embodiments, any VH domain provided herein comprises one or more HC CDR sequences provided herein (e.g., HC CDR1, HC CDR2, and HC CDR3), which are, for example, any of the HC CDR sequences provided for any one of the anti-KLK5 / KLK7 selected from Tables 1a and 1b. In some embodiments, each VL domain provided herein comprises one or more LC CDR sequences provided herein (e.g., LC CDR1, LC CDR2, and LC CDR3), which are, for example, any of the LC CDR sequences provided for any one of the anti-KLK5 / KLK7 antibodies selected from Tables 1a and 1b.

[0083] In some embodiments, the antibodies of the Disclosure include any antibody comprising a heavy chain variable domain and / or light chain variable domain of any one of the anti-KLK5 / KLK7 antibodies selected from Tables 1a and 1b, as well as variants thereof. In some embodiments, the antibodies of the Disclosure include any antibody comprising a pair of heavy chain variable and light chain variable regions of any anti-KLK5 / KLK7 antibody selected from Tables 1a and 1b.

[0084] Aspects of this disclosure provide antibodies comprising a heavy chain variable (VH) domain amino acid sequence and / or a light chain variable (VL) domain amino acid sequence homologous to any of the amino acid sequences described herein. In some embodiments, the antibody comprises a heavy chain variable sequence or a light chain variable sequence that is at least 75% (e.g., at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%) identical to any one of the anti-KLK5 / KLK7 antibodies selected from Tables 1a and 1b. In some embodiments, the homologous heavy chain variable amino acid sequence and / or light chain variable amino acid sequence are not mutated in any of the CDR sequences provided herein. For example, in some embodiments, the degree of sequence variation (e.g., at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%) may occur within the heavy chain variable sequence and / or light chain variable sequence, excluding any of the CDR sequences provided herein. In some embodiments, the antibodies provided herein include a heavy chain variable sequence and a light chain variable sequence that include a framework sequence which is at least 75% (e.g., at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%) identical to the framework sequence of any anti-KLK5 / KLK7 antibody selected from Tables 1a and 1b.

[0085] In some embodiments, the antibody of the Disclosure is a humanized antibody (e.g., a humanized variant containing one or more CDRs from Tables 1a and 1b). In some embodiments, the antibody of the Disclosure comprises HC CDR1, HC CDR2, HC CDR3, LC CDR1, LC CDR2, and LC CDR3, which are the same as HC CDR1, HC CDR2, HC CDR3, LC CDR1, LC CDR2, and LC CDR3 shown in Tables 1a and 1b, and includes a humanized heavy chain variable region and / or a humanized light chain variable region.

[0086] In some embodiments, the antibody of the Disclosure is a humanized antibody comprising a VH containing 20 or fewer amino acid mutations (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid mutations) compared to any VH of any of the anti-KLK5 / KLK7 antibodies listed in Tables 1a and 1b. Alternatively or additionally, the antibody of the Disclosure is a humanized antibody comprising a VL containing 20 or fewer amino acid mutations (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid mutations) compared to any one VL of any of the anti-KLK5 / KLK7 antibodies listed in Tables 1a and 1b.

[0087] In some embodiments, the anti-KLK5 / KLK7 antibody of the present disclosure comprises heavy chain variable domains HC CDR1, HC CDR2, and HC CDR3 having the amino acid sequence of SEQ ID NO: 7. Alternatively or additionally, the anti-KLK5 / KLK7 antibody of the present disclosure comprises light chain variable domains LC CDR1, LC CDR2, and LC CDR3 having the amino acid sequence of SEQ ID NO: 8.

[0088] In some embodiments, the anti-KLK5 / KLK7 antibody of the present disclosure includes HC CDR1 having the amino acid sequence of SEQ ID NO: 1, HC CDR2 having the amino acid sequence of SEQ ID NO: 2, HC CDR3 having the amino acid sequence of SEQ ID NO: 3, LC CDR1 having the amino acid sequence of SEQ ID NO: 4, LC CDR2 having the amino acid sequence of SEQ ID NO: 5, and LC CDR3 having the amino acid sequence of SEQ ID NO: 6.

[0089] In some embodiments, the anti-KLK5 / KLK7 antibodies of this disclosure comprise HC CDR1, HC CDR2, and HC CDR3, which collectively contain 5 or fewer amino acid mutations (e.g., 5, 4, 3, 2, or 1 or fewer amino acid mutations) compared to HC CDR1 having the amino acid sequence of SEQ ID NO: 1, HC CDR2 having the amino acid sequence of SEQ ID NO: 2, and HC CDR3 having the amino acid sequence of SEQ ID NO: 3. As used anywhere in this disclosure, “collectively” means that the total number of amino acid mutations in all three heavy chain CDRs is within a defined range. Alternatively or additionally, the anti-KLK5 / KLK7 antibodies of this disclosure comprise LC CDR1, LC CDR2, and LC CDR3, which collectively contain 5 or fewer amino acid mutations (e.g., 5, 4, 3, 2, or 1 or fewer amino acid mutations) compared to LC CDR1 having the amino acid sequence of SEQ ID NO: 4, LC CDR2 having the amino acid sequence of SEQ ID NO: 5, and LC CDR3 having the amino acid sequence of SEQ ID NO: 6.

[0090] In some embodiments, the anti-KLK5 / KLK7 antibody of the present disclosure comprises HC CDR1, HC CDR2, and HC CDR3 which are collectively at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%) identical to HC CDR1 having the amino acid sequence of SEQ ID NO: 1, HC CDR2 having the amino acid sequence of SEQ ID NO: 2, and HC CDR3 having the amino acid sequence of SEQ ID NO: 3. Alternatively or additionally, the anti-KLK5 / KLK7 antibodies of the present disclosure include LC CDR1, LC CDR2, and LC CDR3 which are collectively at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%) identical to LC CDR1 having the amino acid sequence of SEQ ID NO: 4, LC CDR2 having the amino acid sequence of SEQ ID NO: 5, and LC CDR3 having the amino acid sequence of SEQ ID NO: 6.

[0091] In some embodiments, the anti-KLK5 / KLK7 antibody of the present disclosure includes HC CDR1 having 3 or fewer amino acid mutations (e.g., 3, 2, or 1 or fewer amino acid mutations) compared to HC CDR1 having the amino acid sequence of SEQ ID NO: 1; HC CDR2 having 3 or fewer amino acid mutations (e.g., 3, 2, or 1 or fewer amino acid mutations) compared to HC CDR2 having the amino acid sequence of SEQ ID NO: 2; and / or HC CDR3 having 3 or fewer amino acid mutations (e.g., 3, 2, or 1 or fewer amino acid mutations) compared to HC CDR3 having the amino acid sequence of SEQ ID NO: 3. Alternatively or additionally, the anti-KLK5 / KLK7 antibodies of this disclosure include LC CDR1 having 3 or fewer amino acid mutations (e.g., 3, 2, or 1 or fewer amino acid mutations) compared to LC CDR1 having the amino acid sequence of SEQ ID NO: 4; LC CDR2 having 3 or fewer amino acid mutations (e.g., 3, 2, or 1 or fewer amino acid mutations) compared to LC CDR2 having the amino acid sequence of SEQ ID NO: 5; and / or LC CDR3 having 3 or fewer amino acid mutations (e.g., 3, 2, or 1 or fewer amino acid mutations) compared to LC CDR3 having the amino acid sequence of SEQ ID NO: 6.

[0092] In some embodiments, the anti-KLK5 / KLK7 antibody of the present disclosure comprises VH, which contains the amino acid sequence of SEQ ID NO: 7. Alternatively or additionally, the anti-KLK5 / KLK7 antibody of the present disclosure comprises VL, which contains the amino acid sequence of SEQ ID NO: 8.

[0093] In some embodiments, the anti-KLK5 / KLK7 antibody of this disclosure includes a VH containing 20 or fewer amino acid mutations (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid mutations) compared to the VH described in SEQ ID NO: 7. Alternatively or additionally, the anti-KLK5 / KLK7 antibody of this disclosure includes a VL containing 20 or fewer amino acid mutations (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid mutations) compared to the VL described in SEQ ID NO: 8.

[0094] In some embodiments, the anti-KLK5 / KLK7 antibody of the present disclosure includes a VH having an amino acid sequence that is at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%) identical to the VH described in SEQ ID NO: 7. Alternatively or additionally, the anti-KLK5 / KLK7 antibody of the present disclosure includes a VL having an amino acid sequence that is at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%) identical to the VL described in SEQ ID NO: 8.

[0095] In some embodiments, the anti-KLK5 / KLK7 antibody of the present disclosure comprises heavy chain variable domains HC CDR1, HC CDR2, and HC CDR3 having the amino acid sequence of SEQ ID NO: 13. Alternatively or additionally, the anti-KLK5 / KLK7 antibody of the present disclosure comprises light chain variable domains LC CDR1, LC CDR2, and LC CDR3 having the amino acid sequence of SEQ ID NO: 14.

[0096] In some embodiments, the anti-KLK5 / KLK7 antibody of the present disclosure includes HC CDR1 having the amino acid sequence of SEQ ID NO: 9, HC CDR2 having the amino acid sequence of SEQ ID NO: 10, HC CDR3 having the amino acid sequence of SEQ ID NO: 11, LC CDR1 having the amino acid sequence of SEQ ID NO: 4, LC CDR2 having the amino acid sequence of SEQ ID NO: 5, and LC CDR3 having the amino acid sequence of SEQ ID NO: 12.

[0097] In some embodiments, the anti-KLK5 / KLK7 antibodies of this disclosure comprise HC CDR1, HC CDR2, and HC CDR3, which collectively contain 5 or fewer amino acid mutations (e.g., 5, 4, 3, 2, or 1 or fewer amino acid mutations) compared to HC CDR1 having the amino acid sequence of SEQ ID NO: 9, HC CDR2 having the amino acid sequence of SEQ ID NO: 10, and HC CDR3 having the amino acid sequence of SEQ ID NO: 11. As used anywhere in this disclosure, “collectively” means that the total number of amino acid mutations in all three heavy chain CDRs is within a defined range. Alternatively or additionally, the anti-KLK5 / KLK7 antibodies of this disclosure comprise LC CDR1, LC CDR2, and LC CDR3, which collectively contain 5 or fewer amino acid mutations (e.g., 5, 4, 3, 2, or 1 or fewer amino acid mutations) compared to LC CDR1 having the amino acid sequence of SEQ ID NO: 4, LC CDR2 having the amino acid sequence of SEQ ID NO: 5, and LC CDR3 having the amino acid sequence of SEQ ID NO: 12.

[0098] In some embodiments, the anti-KLK5 / KLK7 antibody of the present disclosure comprises HC CDR1, HC CDR2, and HC CDR3 which are collectively at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%) identical to HC CDR1 having the amino acid sequence of SEQ ID NO: 9, HC CDR2 having the amino acid sequence of SEQ ID NO: 10, and HC CDR3 having the amino acid sequence of SEQ ID NO: 11. Alternatively or additionally, the anti-KLK5 / KLK7 antibodies of the present disclosure include LC CDR1, LC CDR2, and LC CDR3 which are collectively at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%) identical to LC CDR1 having the amino acid sequence of SEQ ID NO: 4, LC CDR2 having the amino acid sequence of SEQ ID NO: 5, and LC CDR3 having the amino acid sequence of SEQ ID NO: 12.

[0099] In some embodiments, the anti-KLK5 / KLK7 antibody of the present disclosure includes HC CDR1 having 3 or fewer amino acid mutations (e.g., 3, 2, or 1 or fewer amino acid mutations) compared to HC CDR1 having the amino acid sequence of SEQ ID NO. 9; HC CDR2 having 3 or fewer amino acid mutations (e.g., 3, 2, or 1 or fewer amino acid mutations) compared to HC CDR2 having the amino acid sequence of SEQ ID NO. 10; and / or HC CDR3 having 3 or fewer amino acid mutations (e.g., 3, 2, or 1 or fewer amino acid mutations) compared to HC CDR3 having the amino acid sequence of SEQ ID NO. 11. Alternatively or additionally, the anti-KLK5 / KLK7 antibodies of this disclosure include LC CDR1 having 3 or fewer amino acid mutations (e.g., 3, 2, or 1 or fewer amino acid mutations) compared to LC CDR1 having the amino acid sequence of SEQ ID NO: 4; LC CDR2 having 3 or fewer amino acid mutations (e.g., 3, 2, or 1 or fewer amino acid mutations) compared to LC CDR2 having the amino acid sequence of SEQ ID NO: 5; and / or LC CDR3 having 3 or fewer amino acid mutations (e.g., 3, 2, or 1 or fewer amino acid mutations) compared to LC CDR3 having the amino acid sequence of SEQ ID NO: 12.

[0100] In some embodiments, the anti-KLK5 / KLK7 antibody of the present disclosure comprises VH, which contains the amino acid sequence of SEQ ID NO: 13. Alternatively or additionally, the anti-KLK5 / KLK7 antibody of the present disclosure comprises VL, which contains the amino acid sequence of SEQ ID NO: 14.

[0101] In some embodiments, the anti-KLK5 / KLK7 antibody of this disclosure includes a VH containing 20 or fewer amino acid mutations (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid mutations) compared to the VH described in SEQ ID NO. 13. Alternatively or additionally, the anti-KLK5 / KLK7 antibody of this disclosure includes a VL containing 20 or fewer amino acid mutations (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid mutations) compared to the VL described in SEQ ID NO. 14.

[0102] In some embodiments, the anti-KLK5 / KLK7 antibody of the present disclosure includes a VH having an amino acid sequence that is at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%) identical to the VH described in SEQ ID NO: 13. Alternatively or additionally, the anti-KLK5 / KLK7 antibody of the present disclosure includes a VL having an amino acid sequence that is at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%) identical to the VL described in SEQ ID NO: 14.

[0103] In some embodiments, the anti-KLK5 / KLK7 antibody of the present disclosure comprises heavy chain variable domains HC CDR1, HC CDR2, and HC CDR3 having the amino acid sequence of SEQ ID NO: 17. Alternatively or additionally, the anti-KLK5 / KLK7 antibody of the present disclosure comprises light chain variable domains LC CDR1, LC CDR2, and LC CDR3 having the amino acid sequence of SEQ ID NO: 14.

[0104] In some embodiments, the anti-KLK5 / KLK7 antibody of the present disclosure includes HC CDR1 having the amino acid sequence of SEQ ID NO: 9, HC CDR2 having the amino acid sequence of SEQ ID NO: 15, HC CDR3 having the amino acid sequence of SEQ ID NO: 16, LC CDR1 having the amino acid sequence of SEQ ID NO: 4, LC CDR2 having the amino acid sequence of SEQ ID NO: 5, and LC CDR3 having the amino acid sequence of SEQ ID NO: 12.

[0105] In some embodiments, the anti-KLK5 / KLK7 antibodies of this disclosure comprise HC CDR1, HC CDR2, and HC CDR3, which collectively contain 5 or fewer amino acid mutations (e.g., 5, 4, 3, 2, or 1 or fewer amino acid mutations) compared to HC CDR1 having the amino acid sequence of SEQ ID NO: 9, HC CDR2 having the amino acid sequence of SEQ ID NO: 15, and HC CDR3 having the amino acid sequence of SEQ ID NO: 16. As used anywhere in this disclosure, “collectively” means that the total number of amino acid mutations in all three heavy chain CDRs is within a defined range. Alternatively or additionally, the anti-KLK5 / KLK7 antibodies of this disclosure comprise LC CDR1, LC CDR2, and LC CDR3, which collectively contain 5 or fewer amino acid mutations (e.g., 5, 4, 3, 2, or 1 or fewer amino acid mutations) compared to LC CDR1 having the amino acid sequence of SEQ ID NO: 4, LC CDR2 having the amino acid sequence of SEQ ID NO: 5, and LC CDR3 having the amino acid sequence of SEQ ID NO: 12.

[0106] In some embodiments, the anti-KLK5 / KLK7 antibody of the present disclosure comprises HC CDR1, HC CDR2, and HC CDR3 which are collectively at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%) identical to HC CDR1 having the amino acid sequence of SEQ ID NO: 9, HC CDR2 having the amino acid sequence of SEQ ID NO: 15, and HC CDR3 having the amino acid sequence of SEQ ID NO: 16. Alternatively or additionally, the anti-KLK5 / KLK7 antibodies of the present disclosure include LC CDR1, LC CDR2, and LC CDR3 which are collectively at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%) identical to LC CDR1 having the amino acid sequence of SEQ ID NO: 4, LC CDR2 having the amino acid sequence of SEQ ID NO: 5, and LC CDR3 having the amino acid sequence of SEQ ID NO: 12.

[0107] In some embodiments, the anti-KLK5 / KLK7 antibody of the present disclosure includes HC CDR1 having 3 or fewer amino acid mutations (e.g., 3, 2, or 1 or fewer amino acid mutations) compared to HC CDR1 having the amino acid sequence of SEQ ID NO. 9; HC CDR2 having 3 or fewer amino acid mutations (e.g., 3, 2, or 1 or fewer amino acid mutations) compared to HC CDR2 having the amino acid sequence of SEQ ID NO. 15; and / or HC CDR3 having 3 or fewer amino acid mutations (e.g., 3, 2, or 1 or fewer amino acid mutations) compared to HC CDR3 having the amino acid sequence of SEQ ID NO. 16. Alternatively or additionally, the anti-KLK5 / KLK7 antibodies of this disclosure include LC CDR1 having 3 or fewer amino acid mutations (e.g., 3, 2, or 1 or fewer amino acid mutations) compared to LC CDR1 having the amino acid sequence of SEQ ID NO: 4; LC CDR2 having 3 or fewer amino acid mutations (e.g., 3, 2, or 1 or fewer amino acid mutations) compared to LC CDR2 having the amino acid sequence of SEQ ID NO: 5; and / or LC CDR3 having 3 or fewer amino acid mutations (e.g., 3, 2, or 1 or fewer amino acid mutations) compared to LC CDR3 having the amino acid sequence of SEQ ID NO: 12.

[0108] In some embodiments, the anti-KLK5 / KLK7 antibody of the present disclosure comprises VH, which contains the amino acid sequence of SEQ ID NO: 17. Alternatively or additionally, the anti-KLK5 / KLK7 antibody of the present disclosure comprises VL, which contains the amino acid sequence of SEQ ID NO: 14.

[0109] In some embodiments, the anti-KLK5 / KLK7 antibody of this disclosure includes a VH containing 20 or fewer amino acid mutations (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid mutations) compared to the VH described in SEQ ID NO. 17. Alternatively or additionally, the anti-KLK5 / KLK7 antibody of this disclosure includes a VL containing 20 or fewer amino acid mutations (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid mutations) compared to the VL described in SEQ ID NO. 14.

[0110] In some embodiments, the anti-KLK5 / KLK7 antibody of the present disclosure includes a VH having an amino acid sequence that is at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%) identical to the VH described in SEQ ID NO: 17. Alternatively or additionally, the anti-KLK5 / KLK7 antibody of the present disclosure includes a VL having an amino acid sequence that is at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%) identical to the VL described in SEQ ID NO: 14.

[0111] In some embodiments, the anti-KLK5 / KLK7 antibody of the present disclosure comprises heavy chain variable domains HC CDR1, HC CDR2, and HC CDR3 having the amino acid sequence of SEQ ID NO: 21. Alternatively or additionally, the anti-KLK5 / KLK7 antibody of the present disclosure comprises light chain variable domains LC CDR1, LC CDR2, and LC CDR3 having the amino acid sequence of SEQ ID NO: 14.

[0112] In some embodiments, the anti-KLK5 / KLK7 antibody of the present disclosure includes HC CDR1 having the amino acid sequence of SEQ ID NO: 18, HC CDR2 having the amino acid sequence of SEQ ID NO: 19, HC CDR3 having the amino acid sequence of SEQ ID NO: 20, LC CDR1 having the amino acid sequence of SEQ ID NO: 4, LC CDR2 having the amino acid sequence of SEQ ID NO: 5, and LC CDR3 having the amino acid sequence of SEQ ID NO: 12.

[0113] In some embodiments, the anti-KLK5 / KLK7 antibodies of this disclosure comprise HC CDR1, HC CDR2, and HC CDR3, which collectively contain 5 or fewer amino acid mutations (e.g., 5, 4, 3, 2, or 1 or fewer amino acid mutations) compared to HC CDR1 having the amino acid sequence of SEQ ID NO: 18, HC CDR2 having the amino acid sequence of SEQ ID NO: 19, and HC CDR3 having the amino acid sequence of SEQ ID NO: 20. As used anywhere in this disclosure, “collectively” means that the total number of amino acid mutations in all three heavy chain CDRs is within a defined range. Alternatively or additionally, the anti-KLK5 / KLK7 antibodies of this disclosure comprise LC CDR1, LC CDR2, and LC CDR3, which collectively contain 5 or fewer amino acid mutations (e.g., 5, 4, 3, 2, or 1 or fewer amino acid mutations) compared to LC CDR1 having the amino acid sequence of SEQ ID NO: 4, LC CDR2 having the amino acid sequence of SEQ ID NO: 5, and LC CDR3 having the amino acid sequence of SEQ ID NO: 12.

[0114] In some embodiments, the anti-KLK5 / KLK7 antibody of the present disclosure comprises HC CDR1, HC CDR2, and HC CDR3 which are collectively at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%) identical to HC CDR1 having the amino acid sequence of SEQ ID NO: 18, HC CDR2 having the amino acid sequence of SEQ ID NO: 19, and HC CDR3 having the amino acid sequence of SEQ ID NO: 20. Alternatively or additionally, the anti-KLK5 / KLK7 antibodies of the present disclosure include LC CDR1, LC CDR2, and LC CDR3 which are collectively at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%) identical to LC CDR1 having the amino acid sequence of SEQ ID NO: 4, LC CDR2 having the amino acid sequence of SEQ ID NO: 5, and LC CDR3 having the amino acid sequence of SEQ ID NO: 12.

[0115] In some embodiments, the anti-KLK5 / KLK7 antibody of the present disclosure includes HC CDR1 having 3 or fewer amino acid mutations (e.g., 3, 2, or 1 or fewer amino acid mutations) compared to HC CDR1 having the amino acid sequence of SEQ ID NO. 18; HC CDR2 having 3 or fewer amino acid mutations (e.g., 3, 2, or 1 or fewer amino acid mutations) compared to HC CDR2 having the amino acid sequence of SEQ ID NO. 19; and / or HC CDR3 having 3 or fewer amino acid mutations (e.g., 3, 2, or 1 or fewer amino acid mutations) compared to HC CDR3 having the amino acid sequence of SEQ ID NO. 20. Alternatively or additionally, the anti-KLK5 / KLK7 antibodies of this disclosure include LC CDR1 having 3 or fewer amino acid mutations (e.g., 3, 2, or 1 or fewer amino acid mutations) compared to LC CDR1 having the amino acid sequence of SEQ ID NO: 4; LC CDR2 having 3 or fewer amino acid mutations (e.g., 3, 2, or 1 or fewer amino acid mutations) compared to LC CDR2 having the amino acid sequence of SEQ ID NO: 5; and / or LC CDR3 having 3 or fewer amino acid mutations (e.g., 3, 2, or 1 or fewer amino acid mutations) compared to LC CDR3 having the amino acid sequence of SEQ ID NO: 12.

[0116] In some embodiments, the anti-KLK5 / KLK7 antibody of the present disclosure comprises VH, which contains the amino acid sequence of SEQ ID NO: 21. Alternatively or additionally, the anti-KLK5 / KLK7 antibody of the present disclosure comprises VL, which contains the amino acid sequence of SEQ ID NO: 14.

[0117] In some embodiments, the anti-KLK5 / KLK7 antibody of this disclosure includes a VH containing 20 or fewer amino acid mutations (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid mutations) compared to the VH described in SEQ ID NO. 21. Alternatively or additionally, the anti-KLK5 / KLK7 antibody of this disclosure includes a VL containing 20 or fewer amino acid mutations (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid mutations) compared to the VL described in SEQ ID NO. 14.

[0118] In some embodiments, the anti-KLK5 / KLK7 antibody of the present disclosure includes a VH having an amino acid sequence that is at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%) identical to the VH described in SEQ ID NO: 21. Alternatively or additionally, the anti-KLK5 / KLK7 antibody of the present disclosure includes a VL having an amino acid sequence that is at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%) identical to the VL described in SEQ ID NO: 14.

[0119] The antibodies described herein may be in any antibody form, including, but are not limited to, intact (i.e., full-length) antibodies, their antigen-binding fragments (Fab, F(ab'), F(ab')2, Fv, etc.), single-chain antibodies, bispecific antibodies, or nanobodies. In some embodiments, the anti-KLK5 / KLK7 antibodies described herein are scFv. In some embodiments, the anti-KLK5 / KLK7 antibodies described herein are scFv-Fab (e.g., scFv fused to a portion of the constant region).

[0120] In some embodiments, the anti-KLK5 / KLK7 antibody of this disclosure is a chimeric antibody, which may include a heavy chain constant region and a light chain constant region derived from a human antibody. A chimeric antibody refers to an antibody having a variable region or a portion of a variable region derived from a first species and a constant region derived from a second species. Typically, in these chimeric antibodies, both the light chain and heavy chain variable regions mimic the variable regions of an antibody derived from one species of mammal (e.g., a non-human mammal such as a mouse, rabbit, and rat), while the constant portion is homologous to the sequence in an antibody derived from another mammal, such as a human. In some embodiments, amino acid modifications may be made in the variable region and / or the constant region.

[0121] In some embodiments, the antibodies of this disclosure comprise a VL domain and / or VH domain of any one of the anti-KLK5 / KLK7 antibodies selected from Tables 1a and 1b, and include a constant region comprising the amino acid sequence of the constant region of an IgG, IgE, IgM, IgD, IgA, or IgY immunoglobulin molecule, any class (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2), or any subclass (e.g., IgG2a and IgG2b) immunoglobulin molecule. Non-limiting examples of human constant regions have been described in the Art; see, for example, Kabat EA et al., (1991), cited above.

[0122] In some embodiments, any of the light chains of the anti-KLK5 / KLK7 antibodies described herein may further comprise a light chain constant region (CL), which can be any CL known in the art. In some examples, the CL is a kappa light chain. In other examples, the CL is a lambda light chain. In some embodiments, the CL is a kappa light chain.

[0123] Other antibody heavy chain constant regions and light chain constant regions are well known in the art and are provided, for example, in the IMGT database (www.imgt.org) or www.vbase2.org / vbstat.php, both of which are incorporated herein by reference.

[0124] In some embodiments, conservative mutations may be introduced into the antibody sequence (e.g., CDR or framework sequence) at a position where the residue is unlikely to be involved in interaction with the target antigen (e.g., human or mouse KLK5 and / or human or mouse KLK7), for example, if determined based on the crystal structure. In some embodiments, one, two, or more mutations (e.g., amino acid substitutions) may be introduced into the Fc region (numbered according to the Kabat numbering system (e.g., the EU index in Kabat), for example, the CH2 domain (residues 231-340 of human IgG1) and / or the CH3 domain (residues 341-447 of human IgG1) and / or the hinge region) of the anti-KLK5 / KLK7 antibody described herein to modify one or more functional properties of the antibody, such as serum half-life, complement fixation, Fc receptor binding, and / or antigen-dependent cytotoxicity.

[0125] In some embodiments, one, two, or more mutations (e.g., amino acid substitutions) are introduced into the hinge region of the Fc region (CH1 domain) to modify (e.g., increase or decrease) the number of cysteine ​​residues in the hinge region, as described, for example, in U.S. Patent No. 5,677,425. Modifying the number of cysteine ​​residues in the hinge region of the CH1 domain can, for example, promote light- and heavy-chain assembly, or modify (e.g., increase or decrease) antibody stability, or promote linker conjugation.

[0126] In some embodiments, one, two, or more mutations (e.g., amino acid substitutions) are introduced into the Fc region of the antibodies described herein (numbered according to the Kabat numbering system (e.g., the EU index in Kabat), for example, the CH2 domain (residues 231-340 of human IgG1) and / or the CH3 domain (residues 341-447 of human IgG1) and / or the hinge region) to increase or decrease the affinity of the antibody to an Fc receptor (e.g., an activated Fc receptor) on the surface of effector cells. Mutations in the Fc region of an antibody that decrease or increase the affinity of the antibody to an Fc receptor, and techniques for introducing such mutations into an Fc receptor or fragment thereof, are known to those skilled in the art. Examples of antibody mutations at the Fc receptor that can be made to alter the affinity of an antibody to the Fc receptor are described, for example, in Smith P et al., (2012) PNAS 109:6181-6186, U.S. Patent No. 6,737,056, and International Publications WO02 / 060919, WO98 / 23289, and WO97 / 34631, which are incorporated herein by reference.

[0127] In some embodiments, one, two, or more amino acid mutations (i.e., substitutions, insertions, or deletions) are introduced into the IgG constant domain or its FcRn binding fragment (preferably an Fc or hinge-Fc domain fragment) to modify (e.g., decrease or increase) the half-life of the antibody in vivo. See, for example, International Publications WO02 / 060919, WO98 / 23289, and WO97 / 34631, and U.S. Patents 5,869,046, 6,121,022, 6,277,375, and 6,165,745 for examples of mutations that modify (e.g., decrease or increase) the half-life of the antibody in vivo.

[0128] In some embodiments, one, two, or more amino acid mutations (i.e., substitutions, insertions, or deletions) are introduced into the IgG constant domain or its FcRn binding fragment (preferably an Fc or hinge-Fc domain fragment) to reduce the half-life of the anti-KLK5 / KLK7 antibody in vivo. In some embodiments, one, two, or more amino acid mutations (i.e., substitutions, insertions, or deletions) are introduced into the IgG constant domain or its FcRn binding fragment (preferably an Fc or hinge-Fc domain fragment) to increase the half-life of the antibody in vivo. In some embodiments, the antibody may have one or more amino acid mutations (e.g., substitutions) in a second constant (CH2) domain (residues 231-340 of human IgG1) and / or a third constant (CH3) domain (residues 341-447 of human IgG1), numbered according to the EU index in Kabat (Kabat EA et al., (1991), cited above). In some embodiments, the constant region of IgG1 of the antibodies described herein, numbered according to the EU index as set forth in Kabat, includes a methionine (M) to tyrosine (Y) substitution at position 252, a serine (S) to threonine (T) substitution at position 254, and a threonine (T) to glutamic acid (E) substitution at position 256. See U.S. Patent No. 7,658,921, which is incorporated herein by reference. This type of mutant IgG is called the “YTE variant” and has been shown to exhibit a four-fold increased half-life compared to the wild-type version of the same antibody (see Dall'Acqua WF et al., (2006) J Biol Chem 281:23514-24). In some embodiments, the antibody includes an IgG constant domain containing one, two, three or more amino acid substitutions of amino acid residues at positions 251–257, 285–290, 308–314, 385–389, and 428–436, numbered according to the EU index as found in Kabat.

[0129] In some embodiments, the antibody includes an Fc region that has been modified, for example, by introducing substitutions of M428L and / or N434A, for the purpose of extending the half-life. Non-limiting examples of such Fc variants that affect the circulating half-life are provided in Saunders KO, Conceptual Approaches to Modulating Antibody Effector Functions and Circulation Half-Life, Front Immunol. 2019;10:1296, which is incorporated herein by reference.

[0130] In some embodiments, the effector function(s) of an anti-KLK5 / KLK7 antibody is modified by introducing one, two, or more amino acid substitutions into the IgG constant domain Fc region, for example, by introducing Leu234Ala and Leu235Ala mutations (commonly referred to as LALA mutations). The effector ligand whose affinity is modified can be, for example, the Fc receptor or the C1 component of complement. This technique is described in U.S. Patents 5,624,821 and 5,648,260. In some embodiments, deletion or inactivation of the constant domain (via point mutation or other means) can reduce the binding of the circulating antibody to the Fc receptor, thereby increasing tumor localization. For a description of mutations that delete or inactivate the constant domain and thereby increase tumor localization, see, for example, U.S. Patents 5,585,097 and 8,591,886. In some embodiments, one or more amino acid substitutions may be introduced into the Fc region of the antibodies described herein to remove potential glycosylation sites on the Fc region, thereby reducing Fc receptor binding (see, for example, Shields RL et al., (2001) J Biol Chem 276:6591-604).

[0131] In some embodiments, one or more amino acids in the constant region of the anti-KLK5 / KLK7 antibody described herein can be replaced with different amino acid residues, thereby giving the antibody modified C1q binding and / or reduced or ineffective complement-dependent cell-mediated cytotoxicity (CDC). This technique is described in detail in U.S. Patent No. 6,194,551 (Idusogie et al). In some embodiments, one or more amino acid residues in the N-terminal region of the CH2 domain of the antibody described herein are modified, thereby altering the antibody's ability to immobilize complement. This technique is further described in International Publication No. WO94 / 29351. In some embodiments, the Fc region of the antibody described herein is modified to increase the antibody's ability to mediate antibody-dependent cell-mediated cytotoxicity (ADCC) and / or increase the antibody's affinity for the Fcγ receptor. This technique is further described in International Publication No. WO00 / 42072.

[0132] In some embodiments, the antibody comprises Fc variants containing amino acid substitutions L234A, L235E, and P329G, in which case the numbering follows the EU index. In some embodiments, the antibody comprising the Fc variant exhibits reduced affinity for one or more or each of FcyRJ, FcyRIIA, FcyRIIIA, and Clq compared to the antibody comprising the wild-type human Fc region. Examples of such Fc variants are provided in International Patent Application Publication WO2021 / 055669, title, FC VARIANTS WITH REDUCED EFFECTOR FUNCTION, published March 25, 2021, and U.S. Patent Application Publication US2021-0087271, title, FC VARIANTS WITH REDUCED EFFECTOR FUNCTION, published March 25, 2021, the contents of which are incorporated herein by reference.

[0133] In some embodiments, as described elsewhere herein, variable domain(s) sequences(options) of the heavy and / or light chains of the antibodies provided herein can be used to generate, for example, CDR-implanted, chimeric, humanized, or compound human antibodies or antigen-binding fragments. As will be understood by those skilled in the art, any variant, CDR-implanted, chimeric, humanized, or compound antibody derived from any of the antibodies provided herein may be useful in the compositions and methods described herein, and the variant, CDR-implanted, chimeric, humanized, or compound antibody will maintain the ability to specifically bind to KLK5 and KLK7 such that it has at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, or more binding to KLK5 and KLK7 compared to the original antibody from which it is derived.

[0134] In some embodiments, the antibodies provided herein include mutations that confer desirable properties to the antibody. For example, to avoid potential complications from Fab-arm exchange, which are known to occur with natural IgG4 mAbs, the antibodies provided herein may include a stabilizing "Adair" mutation (Angal S., et al., “A single amino acid substitution abolishes the heterogeneity of chimeric mouse / human (IgG4) antibody,” Mol Immunol 30, 105-108; 1993), in which serine 228 (EU numbering; residue 241 Kabat numbering) is converted to proline, resulting in an IgG1-like hinge sequence. Thus, any of the antibodies may include a stabilizing "Adair" mutation.

[0135] In some embodiments, the antibody is modified, for example, by glycosylation, phosphorylation, SUMOylation, and / or methylation. In some embodiments, the antibody is a glycosylated antibody conjugated to one or more sugar or carbohydrate molecules. In some embodiments, one or more sugar or carbohydrate molecules are conjugated to the antibody via N-glycosylation, O-glycosylation, C-glycosylation, glyciation (GPI anchor attachment), and / or phosphoglycosylation. In some embodiments, one or more sugar or carbohydrate molecules are monosaccharides, disaccharides, oligosaccharides, or glycans. In some embodiments, one or more sugar or carbohydrate molecules are branched oligosaccharides or branched glycans. In some embodiments, one or more sugar or carbohydrate molecules include mannose units, glucose units, N-acetylglucosamine units, N-acetylgalactosamine units, galactose units, fucose units, or phospholipid units. In some embodiments, there are about 1-10, about 1-5, about 5-10, about 1-4, about 1-3, or about 2 sugar molecules. In some embodiments, the glycosylated antibody is completely or partially glycosylated. In some embodiments, the antibody is glycosylated by chemical reaction or enzymatic means. In some embodiments, the antibody is glycosylated in vitro or intracellularly, and optionally, an enzyme in the N or O glycosylation pathway, such as glycosyltransferase, may be deficient. In some embodiments, the antibody is functionalized with sugar molecules or carbohydrate molecules as described in International Patent Application Publication WO2014065661, published May 1, 2014, titled "Modified antibody, antibody-conjugate and process for the preparation thereof".

[0136] In some embodiments, any one of the anti-KLK5 / KLK7 antibodies described herein may include a signal peptide (e.g., an N-terminal signal peptide) in the heavy chain sequence and / or light chain sequence. In some embodiments, the anti-KLK5 / KLK7 antibody described herein includes either one of the VH sequence and VL sequence, either one of the IgG heavy chain sequence and light chain sequence, or either one of the F(ab') heavy chain sequence and light chain sequence described herein, and further includes a signal peptide (e.g., an N-terminal signal peptide).

[0137] (b) IL-4 / IL-4R targeting antibody Conditions associated with barrier dysfunction are characterized by barrier disruption and inflammation mediated by Th2 helper 2 (Th2)-driven inflammation, such as Th2 cytokines and their receptors, e.g., IL-13, IL-13 receptor (IL-13R), IL-4, IL-4R, IL-5, IL-5R, IL-6, IL-6R, IL-9, IL-9R, IL-31, IL-31R, IL-17E (IL-25), and IL-17ER (IL-25R). In some cases, unregulated KLK activity in the epidermis leads to Th2 cell activation and / or related secretion of Th2 cytokines. Elevated Th2 cytokines can further contribute to KLK hyperactivation (Figure 2). Therapeutic antibodies have been developed to suppress Th2 cytokine-mediated inflammation by targeting Th2 cytokines and / or their respective receptors (called Th2-targeted antibodies), e.g., anti-IL-4R antibodies.

[0138] In some embodiments, the antibodies provided herein include at least one antigen-specific binding site that specifically binds to IL-4 or its receptor (e.g., IL-4R). Antibodies targeting IL-4 or its receptor include, but are not limited to, anti-IL-4 antibodies and anti-IL-4R antibodies. Any suitable IL-4 targeted antibody can be used in methods and compositions, including, but not limited to, those listed in Table 2. [Table 2-1] Table 2-2 Table 2-3 Table 2-4

[0139] In some embodiments, the anti-IL-4 antibody includes an antigen-specific binding site for IL-4 or its receptor. In some embodiments, the anti-IL-4 antibody is used in the following publications: US5552304, published September 3, 1996, title: cDNA Clones Coding For Human Protein Exhibiting A Broad Cellular Activity Spectrum (Human Interleukin-4); US5676940, published October 14, 1997, title: Method Of Reducing Immunoglobulin E Responses; US5985280, published November 16, 1999, title: Diagnosis And / Or Therapy Of Tumours Using Monoclonal Antibodies Specific For The Human IL-4 Receptor; US6358509, published March 19, 2002, title: Antibody Antagonists Of Human Interleukin-4; US5863537, published January 26, 1999, title: Humanized Monoclonal Antibodies Against Human Interleukin-4; WO9414975, published July 7, 1994, title: Monoclonal Antibodies Against The Human Interleukin-4 Receptor And Hybridomas Producing The Same; US5928904, published July 27, 1999, title: DNA Encoding Recombinant IL4 Antibodies Useful In Treatment Of IL4 Mediated Disorders; US5597710, published January 28, 1997, title: Humanized Monoclonal Antibodies Against Human Interleukin-4; US5783181, published July 21, 1998, title: Therapeutic Uses Of Fusion Proteins Between Mutant IL4 / IL13 Antagonists And Immunoglobulins;US2002002132, published January 3, 2002, title: Use Of Interleukin-4 Antagonists And Compositions Thereof; US2007274996, published November 29, 2007, title: Antibodies That Bind Interleukin-4 Receptor; US2008241160, published October 2, 2008, title: Human Monoclonal Antibodies Against Human IL-4; US2010297110, published November 25, 2010, title: Antibody Specific For Human IL-4 For The Treatment Of Cancer; US2008160035, published July 3, 2008, title: High Affinity Human Antibodies To Human IL-4 Receptor; US2010226923, published September 9, 2010, title: Antibodies That Bind IL-4 And / Or IL-13 And Their Uses; US2012097565, published April 26, 2012, title: Stabilized Formulations Containing Anti-Interleukin-4 Receptor (IL-4R) Antibodies; US2014056920, published February 27, 2014, title: Methods For Treating Or Preventing Asthma By Administering An IL-4R Antagonist; US2014072583, published March 13, 2014, title: Methods For Treating Atopic Dermatitis By Administering An IL-4R Antagonist; US2016207995, published July 21, 2016, title: Anti-IL-4 Antibodies And Bispecific Antibodies And Uses Thereof; US2016075777, published March 17, 2016, title: Anti-IL-4 / Anti-IL-13 Bispecific Antibody Formulations;US2014356372, published December 4, 2014, title: Methods For Treating Allergy And Enhancing Allergen-Specific Immunotherapy By Administering An IL-4R Inhibitor; US2015017176, published January 15, 2015, title: Methods For Treating Eosinophilic Esophagitis By Administering An IL-4R Inhibitor; US2015225479, published August 13, 2015, title: Anti-IL-4 / Anti-IL-13 Bispecific Antibody / Polyglutamate Formulations; US2017145089, published May 25, 2017, title: Anti-IL4-IL-13 Bispecific Antibodies; US2017281769, published October 5, 2017, title: Stable Anti-IL-4Ra Formulation; US2018346580, published December 6, 2018, title: Antibodies To Canine Interleukin-4 Receptor Alpha; US2019177408, published June 13, 2019, title: Antibody For Binding To Interleukin 4 Receptor; US2022081485, published March 17, 2022, title: Anti-IL-4R Antibody And Use Thereof; WO19148405, published August 8, 2019, title: IL-4R Antibody And Use Thereof; US2022073631, published March 10, 2022, title: Monoclonal Antibody Against Human Interleukin-4 Receptor Alpha and Use Thereof; US2021238294, published August 5, 2021, title: Human IL-4R Binding Antibody, Antigen Binding Fragment Thereof, And Medical Use Thereof;US2021206861, published July 8, 2021, title: Antibodies Binding To Human IL-4R, Preparation Method Therefor And Use Thereof; US2021403580, published December 30, 2021, title: Human Antibody Having High Affinity To Human IL-4 Receptor Alpha, And Use Thereof; US2023295312, published September 21, 2023, title: Antibody Against Human IL-4Ra And Use Thereof; US2022162328, published May 26, 2022, title: Interleukin-4 Receptor Antibody And Application Thereof; US2022348666, published November 3, 2022, title: Liquid Composition Comprising Antibody Of Human Interleukin-4 Receptor Alpha; US2022411519, published December 29, 2022, title: Anti-IL-4R Single-Domain Antibody And Use Thereof; US2023053131, published February 16, 2023, title: Antibodies To Canine Interleukin-4 Receptor Alpha; US2023088052, published March 23, 2023, title: Pharmaceutical Composition Containing Anti-IL-4R Antibody And Use Thereof; US2023105029, published April 6, 2023, title: Antibodies Binding IL4R And Uses Thereof; US2024067738, published February 29, 2024, title: Anti-IL4 Receptor Antibodies For Veterinary Use;US2023203172, published June 29, 2023, title: Anti-Human Interleukin-4 Receptor Alpha Antibody And Preparation Method And Application Thereof; US2023167180, published June 1, 2023, title: Antibody Binding With Specific Epitope In Human IL-4R Alpha And Applications Of Antibody; WO21254221, published December 23, 2021, title: Stable Liquid Preparation Of Anti-IL-4R Monoclonal Antibodies; US2023279124, published September 7, 2023, title: Fusion Protein Comprising Ige Fc Receptor Alpha Subunit Extracellular Domain And Anti-IL-4R Antibody, And Use Thereof; US2023357415, published November 9, 2023, title: Bispecific Antibody Simultaneously Binding To Interleukin-4 Receptor Alpha Subunit And Interleukin-5 Receptor Alpha Subunit, And Use Thereof; US2023374144, published November 23, 2023, title: Antibodies Specifically Recognizing Interleukin-4 Receptor Alpha And Uses Thereof; US2023279123, published September 7, 2023, title: Humanized Anti-IL-4Ra Single Domain Antibody And Application Thereof; US2024075158, published March 7, 2024, title: Complex Of Anti-IL-4R Antibody Or Antigen-Binding Fragment Thereof And Medical Use Thereof;WO23011502, published February 9, 2023, title: Stable Formulation Comprising Anti-IL-4R Antibody; US2023064378, published March 2, 2023, title: Human Interleukin-4 Receptor Alpha Antibodies; WO23025217, published March 2, 2023, title: Pharmaceutical Composition Of Anti-IL4R Antibody And Use Thereof; WO23191665, published October 5, 2023, title: Antibodies To Human IL-4R Having; The descriptions of any known anti-IL-4 or anti-IL-4R antibodies, including antigen-specific binding sites for IL-4 or IL-4R derived from any known anti-IL-4 or anti-IL-4R antibody, as described in *Reduced Immunogenicity And Application Thereof* (US2024024472, published January 25, 2024, title: Anti-Interleukin-4 Receptor (IL-4R) Antibody Formulations* (US2024100176, published March 28, 2024, title: Human Interleukin-4 Receptor Alpha Antibody Glucocorticoid Conjugates*) and *WO24061279, published March 28, 2024, title: Recombinant Bispecific Antibodies Targeting TSLP And IL4R*, are incorporated herein by reference.

[0140] In some embodiments, the IL-4 targeted antibody includes an antigen-specific binding site for IL-4 or its receptor. In some embodiments, the IL-4 targeted antibody includes HC CDR1, HC CDR2, HC CDR3 of the VH of either the anti-IL-4 antibody or anti-IL-4R antibody from Table 2, and LC CDR1, LC CDR2, and LC CDR3 of the VL of either the anti-IL-4 antibody or anti-IL-4R antibody from Table 2. In some embodiments, the IL-4 targeted antibody includes the VH of either the anti-IL-4 antibody or anti-IL-4R antibody from Table 2, and the VL of either the anti-IL-4 antibody or anti-IL-4R antibody from Table 2. In some embodiments, the IL-4 targeted antibody includes HC CDR1, HC CDR2, HC CDR3 of dupilumab, and LC CDR1, LC CDR2, and LC CDR3 of dupilumab. In some embodiments, the IL-4 targeted antibody includes dupilumab VH and dupilumab VL. In some embodiments, the IL-4 targeted antibody includes pascolizumab HC CDR1, HC CDR2, HC CDR3, and pascolizumab LC CDR1, LC CDR2, and LC CDR3. In some embodiments, the IL-4 targeted antibody includes pascolizumab VH and pascolizumab VL. In some embodiments, the IL-4 targeted antibody includes AMG317 HC CDR1, HC CDR2, HC CDR3, and AMG317 LC CDR1, LC CDR2, and LC CDR3. In some embodiments, the IL-4 targeted antibody includes AMG317 VH and AMG17 ​​VL. In some embodiments, the IL-4 targeted antibody includes HC CDR1, HC CDR2, HC CDR3 of Stapoquivert, and LC CDR1, LC CDR2, and LC CDR3 of Stapoquivert. In some embodiments, the IL-4 targeted antibody includes VH of Stapoquivert and VL of Stapoquivert. In some embodiments, the IL-4 targeted antibody includes HC CDR1, HC CDR2, HC CDR3 of Rademikvert, and LC CDR1, LC CDR2, and LC CDR3 of Rademikvert. In some embodiments, the IL-4 targeted antibody includes VH of Rademikvert and VL of Rademikvert.In some embodiments, the IL-4 targeted antibody includes manfidokimab HC CDR1, HC CDR2, HC CDR3, and manfidokimab LC CDR1, LC CDR2, and LC CDR3. In some embodiments, the IL-4 targeted antibody includes manfidokimab VH and manfidokimab VL.

[0141] (c) Multispecific antibody In some embodiments, the antibodies provided herein are multispecific antibodies (e.g., bispecific antibodies). For example, in some embodiments, one or more anti-KLK5 / KLK7 antibodies may be combined with one or more different anti-KLK5 / KLK7 antibodies to produce multispecific or bispecific anti-KLK5 / KLK7 antibodies. For example, one or more anti-KLK5 / KLK7 antibodies described herein (Tables 1a and 1b) can be combined with one or more different anti-KLK5 / KLK7 antibodies described herein (Tables 1a and 1b) to produce multispecific antibodies.

[0142] In some embodiments, one or more anti-KLK5 / KLK7 antibodies may be combined with any other suitable therapeutic antibody to generate multispecific or bispecific anti-KLK5 / KLK7 / additional target antibodies. For example, the anti-KLK5 / KLK7 antibodies described herein (Tables 1a and 1b) can be combined with any suitable antibody (e.g., the IL-4 targeted antibodies described in Table 2) to generate bispecific antibodies. Such additional therapeutic antibodies include, but are not limited to, anti-IL4R antibodies (e.g., dupilumab).

[0143] Table 6 below shows possible arm configurations for anti-KLK5 / KLK7+IL-4 bispecific antibodies. In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody includes one arm comprising a CDR and / or VH and / or VL, the sequence of which is derived from, corresponds to, or is the same as, the CDR and / or VH and / or VL sequences of the antibodies listed in the left column of Table 6; and a second arm comprising a CDR and / or VH / VL, the sequence of which is derived from, corresponds to, or is the same as, the CDR and / or VH and / or VL sequences of the antibodies listed in the corresponding rows of the right column of Table 6 or another suitable antibody disclosed herein. The CDR and VH / VL sequences of each antibody in Table 6 are listed in Tables 1a and 2. [Table 6]

[0144] In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody comprises one arm containing a CDR and / or VH / VL whose sequence is derived from, corresponds to, or is identical to, the sequence of the CDR and / or VH / VL of an anti-KLK5 / KLK7 antibody listed in Table 1a, and a second arm containing a CDR and / or VH / VL whose sequence is derived from, corresponds to, or is identical to, the sequence of the CDR and / or VH / VL of an anti-IL-4 or anti-IL4R antibody listed in Table 2, Table 6, and elsewhere in this specification. In some embodiments, an anti-KLK5 / KLK7+Th2 bispecific antibody may comprise one arm containing an antigen-specific binding site that targets KLK5 and KLK7, and a second arm containing an antigen-specific binding site that does not target IL-4 or IL-4R.

[0145] In some embodiments, the Disclosure provides a bispecific antibody comprising at least one antigen-specific binding site that specifically binds to KLK5 and KLK7, and at least one antigen-specific binding site that specifically binds to IL-4 or its receptor. In some embodiments, the bispecific antibody described herein comprises at least one antigen-specific binding site that specifically binds to KLK5 and KLK7 (e.g., KLK5 / KLK7-dual-Ab4) and at least one antigen-specific binding site that specifically binds to IL-4 or IL-4R (e.g., one of the anti-IL-4 or anti-IL-4 receptor antibodies described herein). Such a bispecific antibody is referred to as an anti-KLK5 / KLK7+IL-4 bispecific antibody. Such an antigen-specific binding site may be one of the anti-KLK5 / KLK7 antibody and IL-4 / IL-4R antibody disclosed herein.

[0146] In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody described herein includes at least one antigen-specific binding site that specifically binds to KLK5 and KLK7 and binds to the active sites of KLK5 and KLK7. In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody described herein includes at least one antigen-specific binding site that specifically binds to KLK5 and KLK7, which are binding sites of dual inhibitory antibodies that specifically bind to KLK5 and KLK7 (e.g., any one of KLK5 / KLK7-dual-Ab1, KLK5 / KLK7-dual-Ab2, KLK5 / KLK7-dual-Ab3, or KLK5 / KLK7-dual-Ab4). In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody comprises one arm containing at least one antigen-specific binding site that specifically binds to KLK5 and KLK7, including one of the antibodies listed in Tables 1a and 1b (e.g., one of KLK5 / KLK7-dual-Ab1, KLK5 / KLK7-dual-Ab2, KLK5 / KLK7-dual-Ab3, KLK5 / KLK7-dual-Ab4), and one of the anti-IL-4 or anti-IL-4R antibodies listed in Table 2, including one arm containing at least one antigen-specific binding site that specifically binds to KLK5 and KLK7, arm containing at least one antigen-specific binding site that specifically binds to KLK5 and KLK7, including one of the antibodies listed in Tables 1a and 1b (e.g., one of KLK5 / KLK7-dual-Ab1, KLK5 / KLK7-dual-Ab2, KLK5 / KLK7-dual-Ab3, KLK5 / KLK7-dual-Ab4), and one arm containing at least one antigen-specific binding site that specifically binds to KLK5 and KLK7, including one of the anti-IL-4 or anti-IL-4R antibodies listed in Table 2, including one of the anti-IL-4 or anti-IL-4R antibodies, including one arm containing at least one antigen-specific binding site that specifically binds to KLK5 and KLK7, and one arm containing at least one antigen-specific binding site that specifically binds to KLK5 and KLK7, including one of the anti-IL-4 or anti-IL-4R antibodies, including one of the anti-IL-4 or The antibody comprises one arm containing CDR3 and at least one antigen-specific binding site that specifically binds to IL-4 or IL-4R. In some embodiments, the anti-KLK5 / KLK7-IL-4 bispecific antibody comprises one of the antibodies listed in Tables 1a and 1b (e.g., one of KLK5 / KLK7-dual-Ab1, KLK5 / KLK7-dual-Ab2, KLK5 / KLK7-dual-Ab3, KLK5 / KLK7-dual-Ab4) with at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%).One arm containing at least one antigen-specific binding site that specifically binds to KLK5 and KLK7, including a VH that is identical to (or 100%) any one of the antibodies listed in Tables 1a and 1b (e.g., any one of KLK5 / KLK7-dual-Ab1, KLK5 / KLK7-dual-Ab2, KLK5 / KLK7-dual-Ab3, KLK5 / KLK7-dual-Ab4) and a VL that is identical to at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%) any one of the antibodies listed in Tables 1a and 1b (e.g., any one of KLK5 / KLK7-dual-Ab1, KLK5 / KLK7-dual-Ab2, KLK5 / KLK7-dual-Ab3, KLK5 / KLK7-dual-Ab4), and one VH that is identical to at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%) any one of the antibodies listed in Table 2, and at least one VH that is identical to any one of the anti-IL-4 or anti-IL-4R antibodies listed in Table 2 It includes a VH that is at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%) identical, and / or an arm containing at least one antigen-specific binding site that specifically binds to IL-4 or IL-4R, including a VL that is at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100%) identical to one VL of any of the anti-IL-4 or anti-IL-4R antibodies listed in Table 2.

[0147] In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody containing at least one antigen-specific binding site that specifically binds to KLK5 and KLK7 includes heavy chain variable domains HC CDR1, HC CDR2, and HC CDR3 containing the amino acid sequence of SEQ ID NO: 7, and light chain variable domains LC CDR1, LC CDR2, and LC CDR3 containing the amino acid sequence of SEQ ID NO: 8. In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody containing at least one antigen-specific binding site that specifically binds to KLK5 and KLK7 includes heavy chain variable domains HC CDR1, HC CDR2, and HC CDR3 containing the amino acid sequence of SEQ ID NO: 13, and light chain variable domains LC CDR1, LC CDR2, and LC CDR3 containing the amino acid sequence of SEQ ID NO: 14. In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody containing at least one antigen-specific binding site that specifically binds to KLK5 and KLK7 includes heavy chain variable domains HC CDR1, HC CDR2, and HC CDR3 containing the amino acid sequence of SEQ ID NO: 17, and light chain variable domains LC CDR1, LC CDR2, and LC CDR3 containing the amino acid sequence of SEQ ID NO: 14. In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody containing at least one antigen-specific binding site that specifically binds to KLK5 and KLK7 includes heavy chain variable domains HC CDR1, HC CDR2, and HC CDR3 containing the amino acid sequence of SEQ ID NO: 21, and light chain variable domains LC CDR1, LC CDR2, and LC CDR3 containing the amino acid sequence of SEQ ID NO: 14.

[0148] In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody comprising at least one antigen-specific binding site that specifically binds to KLK5 and KLK7 comprises HC CDR1 comprising the amino acid sequence of SEQ ID NO: 1, HC CDR2 comprising the amino acid sequence of SEQ ID NO: 2, HC CDR3 comprising the amino acid sequence of SEQ ID NO: 3, LC CDR1 comprising the amino acid sequence of SEQ ID NO: 4, LC CDR2 comprising the amino acid sequence of SEQ ID NO: 5, and LC CDR3 comprising the amino acid sequence of SEQ ID NO: 6. In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody comprising at least one antigen-specific binding site that specifically binds to KLK5 and KLK7 comprises HC CDR1 comprising the amino acid sequence of SEQ ID NO: 9, HC CDR2 comprising the amino acid sequence of SEQ ID NO: 10, HC CDR3 comprising the amino acid sequence of SEQ ID NO: 11, LC CDR1 comprising the amino acid sequence of SEQ ID NO: 4, LC CDR2 comprising the amino acid sequence of SEQ ID NO: 5, and LC CDR3 comprising the amino acid sequence of SEQ ID NO: 12. In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody comprising at least one antigen-specific binding site that specifically binds to KLK5 and KLK7 includes HC CDR1 comprising the amino acid sequence of SEQ ID NO: 9, HC CDR2 comprising the amino acid sequence of SEQ ID NO: 15, HC CDR3 comprising the amino acid sequence of SEQ ID NO: 16, LC CDR1 comprising the amino acid sequence of SEQ ID NO: 4, LC CDR2 comprising the amino acid sequence of SEQ ID NO: 5, and LC CDR3 comprising the amino acid sequence of SEQ ID NO: 12. In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody comprising at least one antigen-specific binding site that specifically binds to KLK5 and KLK7 includes HC CDR1 comprising the amino acid sequence of SEQ ID NO: 18, HC CDR2 comprising the amino acid sequence of SEQ ID NO: 19, HC CDR3 comprising the amino acid sequence of SEQ ID NO: 20, LC CDR1 comprising the amino acid sequence of SEQ ID NO: 4, LC CDR2 comprising the amino acid sequence of SEQ ID NO: 5, and LC CDR3 comprising the amino acid sequence of SEQ ID NO: 12.

[0149] In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody containing at least one antigen-specific binding site that specifically binds to KLK5 and KLK7 includes a VH containing the amino acid sequence of SEQ ID NO: 7 and a VL containing the amino acid sequence of SEQ ID NO: 8. In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody containing at least one antigen-specific binding site that specifically binds to KLK5 and KLK7 includes a VH containing the amino acid sequence of SEQ ID NO: 13 and a VL containing the amino acid sequence of SEQ ID NO: 14. In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody containing at least one antigen-specific binding site that specifically binds to KLK5 and KLK7 includes a VH containing the amino acid sequence of SEQ ID NO: 17 and a VL containing the amino acid sequence of SEQ ID NO: 14. In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody containing at least one antigen-specific binding site that specifically binds to KLK5 and KLK7 includes VH containing the amino acid sequence of SEQ ID NO: 21 and VL containing the amino acid sequence of SEQ ID NO: 14.

[0150] In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody includes at least one antigen-specific binding site that specifically binds to IL-4 or IL-4R. In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody including at least one antigen-specific binding site that specifically binds to IL-4 or IL-4R includes one of the anti-IL-4 antibodies or anti-IL-4R antibodies listed in Table 2: HC CDR1, HC CDR2, HC CDR3, LC CDR1, LC CDR2, and / or LC CDR3. In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody including at least one antigen-specific binding site that specifically binds to IL-4 or IL-4R includes one of the anti-IL-4 antibodies or anti-IL-4R antibodies listed in Table 2: VH and / or VL.

[0151] In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody includes at least one antigen-specific binding site that specifically binds to KLK5 / KLK7 derived from any one of KLK5 / KLK7-dual-Ab1, KLK5 / KLK7-dual-Ab2, KLK5 / KLK7-dual-Ab3, and KLK5 / KLK7-dual-Ab4, and at least one antigen-specific binding site that specifically binds to IL-4R derived from dupilumab. In some embodiments, the KLK5 / KLK7-IL-4 bispecific antibody, which includes at least one antigen-specific binding site that specifically binds to IL-4R, includes heavy chain variable domains HC CDR1, HC CDR2, and HC CDR3 containing the amino acid sequence of SEQ ID NO: 154, and light chain variable domains LC CDR1, LC CDR2, and LC CDR3 containing the amino acid sequence of SEQ ID NO: 155.

[0152] In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody comprising at least one antigen-specific binding site that specifically binds to IL-4R includes HC CDR1 comprising the amino acid sequence of SEQ ID NO: 156, HC CDR2 comprising the amino acid sequence of SEQ ID NO: 157, HC CDR3 comprising the amino acid sequence of SEQ ID NO: 158, LC CDR1 comprising the amino acid sequence of SEQ ID NO: 159, LC CDR2 comprising the amino acid sequence of SEQ ID NO: 160, and LC CDR3 comprising the amino acid sequence of SEQ ID NO: 161. In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody comprising at least one antigen-specific binding site that specifically binds to IL-4R includes HC CDR1 comprising the amino acid sequence of SEQ ID NO: 162, HC CDR2 comprising the amino acid sequence of SEQ ID NO: 163, HC CDR3 comprising the amino acid sequence of SEQ ID NO: 164, LC CDR1 comprising the amino acid sequence of SEQ ID NO: 165, LC CDR2 comprising the amino acid sequence of LGS, and LC CDR3 comprising the amino acid sequence of SEQ ID NO: 167. In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody, which includes at least one antigen-specific binding site that specifically binds to IL-4R, comprises HC CDR1 containing the amino acid sequence of SEQ ID NO: 168, HC CDR2 containing the amino acid sequence of SEQ ID NO: 169, HC CDR3 containing the amino acid sequence of SEQ ID NO: 170, LC CDR1 containing the amino acid sequence of SEQ ID NO: 171, LC CDR2 containing the amino acid sequence of SEQ ID NO: 172, and LC CDR3 containing the amino acid sequence of SEQ ID NO: 173. In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody, which includes at least one antigen-specific binding site that specifically binds to IL-4R, comprises VH containing the amino acid sequence of SEQ ID NO: 154, and VL containing the amino acid sequence of SEQ ID NO: 155.

[0153] In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody includes at least one antigen-specific binding site that specifically binds to KLK5 / KLK7 derived from any one of KLK5 / KLK7-dual-Ab1, KLK5 / KLK7-dual-Ab2, KLK5 / KLK7-dual-Ab3, and KLK5 / KLK7-dual-Ab4, and at least one antigen-specific binding site that specifically binds to IL-4 derived from pascolizumab. In some embodiments, the KLK5 / KLK7-IL-4 bispecific antibody, which includes at least one antigen-specific binding site that specifically binds to IL-4R, includes heavy chain variable domains HC CDR1, HC CDR2, and HC CDR3 containing the amino acid sequence of SEQ ID NO: 174, and light chain variable domains LC CDR1, LC CDR2, and LC CDR3 containing the amino acid sequence of SEQ ID NO: 175.

[0154] In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody containing at least one antigen-specific binding site that specifically binds to IL-4 includes HC CDR1 containing the amino acid sequence of SEQ ID NO: 176, HC CDR2 containing the amino acid sequence of SEQ ID NO: 177, HC CDR3 containing the amino acid sequence of SEQ ID NO: 178, LC CDR1 containing the amino acid sequence of SEQ ID NO: 179, LC CDR2 containing the amino acid sequence of SEQ ID NO: 180, and LC CDR3 containing the amino acid sequence of SEQ ID NO: 181. In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody containing at least one antigen-specific binding site that specifically binds to IL-4 includes HC CDR1 containing the amino acid sequence of SEQ ID NO: 182, HC CDR2 containing the amino acid sequence of SEQ ID NO: 183, HC CDR3 containing the amino acid sequence of SEQ ID NO: 184, LC CDR1 containing the amino acid sequence of SEQ ID NO: 185, LC CDR2 containing the amino acid sequence of AAS, and LC CDR3 containing the amino acid sequence of SEQ ID NO: 187. In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody, which includes at least one antigen-specific binding site that specifically binds to IL-4, comprises HC CDR1 containing the amino acid sequence of SEQ ID NO: 188, HC CDR2 containing the amino acid sequence of SEQ ID NO: 189, HC CDR3 containing the amino acid sequence of SEQ ID NO: 190, LC CDR1 containing the amino acid sequence of SEQ ID NO: 191, LC CDR2 containing the amino acid sequence of SEQ ID NO: 192, and LC CDR3 containing the amino acid sequence of SEQ ID NO: 193. In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody, which includes at least one antigen-specific binding site that specifically binds to IL-4, comprises VH containing the amino acid sequence of SEQ ID NO: 174, and VL containing the amino acid sequence of SEQ ID NO: 175.

[0155] In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody includes at least one antigen-specific binding site that specifically binds to KLK5 / KLK7 derived from any one of KLK5 / KLK7-dual-Ab1, KLK5 / KLK7-dual-Ab2, KLK5 / KLK7-dual-Ab3, and KLK5 / KLK7-dual-Ab4, and at least one antigen-specific binding site that specifically binds to IL-4R derived from AMG317. In some embodiments, the anti-KLK5 / KLK7-IL-4 bispecific antibody, which includes at least one antigen-specific binding site that specifically binds to IL-4R, includes heavy chain variable domains HC CDR1, HC CDR2, and HC CDR3 containing the amino acid sequence of SEQ ID NO: 194, and light chain variable domains LC CDR1, LC CDR2, and LC CDR3 containing the amino acid sequence of SEQ ID NO: 195.

[0156] In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody containing at least one antigen-specific binding site that specifically binds to IL-4R includes HC CDR1 containing the amino acid sequence of SEQ ID NO: 196, HC CDR2 containing the amino acid sequence of SEQ ID NO: 197, HC CDR3 containing the amino acid sequence of SEQ ID NO: 198, LC CDR1 containing the amino acid sequence of SEQ ID NO: 199, LC CDR2 containing the amino acid sequence of SEQ ID NO: 200, and LC CDR3 containing the amino acid sequence of SEQ ID NO: 201. In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody containing at least one antigen-specific binding site that specifically binds to IL-4R includes HC CDR1 containing the amino acid sequence of SEQ ID NO: 202, HC CDR2 containing the amino acid sequence of SEQ ID NO: 203, HC CDR3 containing the amino acid sequence of SEQ ID NO: 204, LC CDR1 containing the amino acid sequence of SEQ ID NO: 205, LC CDR2 containing the amino acid sequence of SEQ ID NO: 206, and LC CDR3 containing the amino acid sequence of SEQ ID NO: 207. In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody, which includes at least one antigen-specific binding site that specifically binds to IL-4R, includes HC CDR1 containing the amino acid sequence of SEQ ID NO: 208, HC CDR2 containing the amino acid sequence of SEQ ID NO: 209, HC CDR3 containing the amino acid sequence of SEQ ID NO: 210, LC CDR1 containing the amino acid sequence of SEQ ID NO: 211, LC CDR2 containing the amino acid sequence of GAS, and LC CDR3 containing the amino acid sequence of SEQ ID NO: 213. In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody, which includes at least one antigen-specific binding site that specifically binds to IL-4R, includes VH containing the amino acid sequence of SEQ ID NO: 194, and VL containing the amino acid sequence of SEQ ID NO: 195.

[0157] In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody includes at least one antigen-specific binding site that specifically binds to KLK5 / KLK7 derived from any one of KLK5 / KLK7-dual-Ab1, KLK5 / KLK7-dual-Ab2, KLK5 / KLK7-dual-Ab3, and KLK5 / KLK7-dual-Ab4, and at least one antigen-specific binding site that specifically binds to IL-4 derived from stabokivart. In some embodiments, the anti-KLK5 / KLK7-IL-4R bispecific antibody, which includes at least one antigen-specific binding site that specifically binds to IL-4R, includes heavy chain variable domains HC CDR1, HC CDR2, and HC CDR3 containing the amino acid sequence of SEQ ID NO: 214, and light chain variable domains LC CDR1, LC CDR2, and LC CDR3 containing the amino acid sequence of SEQ ID NO: 215.

[0158] In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody comprising at least one antigen-specific binding site that specifically binds to IL-4R includes HC CDR1 comprising the amino acid sequence of SEQ ID NO: 216, HC CDR2 comprising the amino acid sequence of SEQ ID NO: 217, HC CDR3 comprising the amino acid sequence of SEQ ID NO: 218, LC CDR1 comprising the amino acid sequence of SEQ ID NO: 219, LC CDR2 comprising the amino acid sequence of SEQ ID NO: 220, and LC CDR3 comprising the amino acid sequence of SEQ ID NO: 221. In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody comprising at least one antigen-specific binding site that specifically binds to IL-4R includes HC CDR1 comprising the amino acid sequence of SEQ ID NO: 222, HC CDR2 comprising the amino acid sequence of SEQ ID NO: 223, HC CDR3 comprising the amino acid sequence of SEQ ID NO: 224, LC CDR1 comprising the amino acid sequence of SEQ ID NO: 225, LC CDR2 comprising the amino acid sequence of YTS, and LC CDR3 comprising the amino acid sequence of SEQ ID NO: 227. In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody, comprising at least one antigen-specific binding site that specifically binds to IL-4R, includes HC CDR1 comprising the amino acid sequence of SEQ ID NO: 228, HC CDR2 comprising the amino acid sequence of SEQ ID NO: 229, HC CDR3 comprising the amino acid sequence of SEQ ID NO: 230, LC CDR1 comprising the amino acid sequence of SEQ ID NO: 231, LC CDR2 comprising the amino acid sequence of SEQ ID NO: 232, and LC CDR3 comprising the amino acid sequence of SEQ ID NO: 233. In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody, comprising at least one antigen-specific binding site that specifically binds to IL-4R, includes VH comprising the amino acid sequence of SEQ ID NO: 214, and VL comprising the amino acid sequence of SEQ ID NO: 215.

[0159] In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody includes at least one antigen-specific binding site that specifically binds to KLK5 / KLK7 derived from any one of KLK5 / KLK7-dual-Ab1, KLK5 / KLK7-dual-Ab2, KLK5 / KLK7-dual-Ab3, and KLK5 / KLK7-dual-Ab4, and at least one antigen-specific binding site that specifically binds to IL-4R derived from rademiquivert. In some embodiments, the anti-KLK5 / KLK7-IL-4 bispecific antibody, which includes at least one antigen-specific binding site that specifically binds to IL-4R, includes heavy chain variable domains HC CDR1, HC CDR2, and HC CDR3 containing the amino acid sequence of SEQ ID NO: 234, and light chain variable domains LC CDR1, LC CDR2, and LC CDR3 containing the amino acid sequence of SEQ ID NO: 235.

[0160] In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody comprising at least one antigen-specific binding site that specifically binds to IL-4R includes HC CDR1 comprising the amino acid sequence of SEQ ID NO: 236, HC CDR2 comprising the amino acid sequence of SEQ ID NO: 237, HC CDR3 comprising the amino acid sequence of SEQ ID NO: 238, LC CDR1 comprising the amino acid sequence of SEQ ID NO: 239, LC CDR2 comprising the amino acid sequence of SEQ ID NO: 240, and LC CDR3 comprising the amino acid sequence of SEQ ID NO: 241. In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody comprising at least one antigen-specific binding site that specifically binds to IL-4R includes HC CDR1 comprising the amino acid sequence of SEQ ID NO: 242, HC CDR2 comprising the amino acid sequence of SEQ ID NO: 243, HC CDR3 comprising the amino acid sequence of SEQ ID NO: 244, LC CDR1 comprising the amino acid sequence of SEQ ID NO: 245, LC CDR2 comprising the amino acid sequence of SEQ ID NO: 246, and LC CDR3 comprising the amino acid sequence of SEQ ID NO: 247. In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody, which includes at least one antigen-specific binding site that specifically binds to IL-4R, includes HC CDR1 containing the amino acid sequence of SEQ ID NO: 248, HC CDR2 containing the amino acid sequence of SEQ ID NO: 249, HC CDR3 containing the amino acid sequence of SEQ ID NO: 250, LC CDR1 containing the amino acid sequence of SEQ ID NO: 251, LC CDR2 containing the amino acid sequence of GAS, and LC CDR3 containing the amino acid sequence of SEQ ID NO: 253. In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody, which includes at least one antigen-specific binding site that specifically binds to IL-4R, includes VH containing the amino acid sequence of SEQ ID NO: 234, and VL containing the amino acid sequence of SEQ ID NO: 235.

[0161] In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody includes at least one antigen-specific binding site that specifically binds to KLK5 / KLK7 derived from any one of KLK5 / KLK7-dual-Ab1, KLK5 / KLK7-dual-Ab2, KLK5 / KLK7-dual-Ab3, and KLK5 / KLK7-dual-Ab4, and at least one antigen-specific binding site that specifically binds to IL-4R derived from manfidokimab. In some embodiments, the anti-KLK5 / KLK7-IL-4 bispecific antibody, which includes at least one antigen-specific binding site that specifically binds to IL-4R, includes heavy chain variable domains HC CDR1, HC CDR2, and HC CDR3 containing the amino acid sequence of SEQ ID NO: 254, and light chain variable domains LC CDR1, LC CDR2, and LC CDR3 containing the amino acid sequence of SEQ ID NO: 255.

[0162] In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody comprising at least one antigen-specific binding site that specifically binds to IL-4R includes HC CDR1 comprising the amino acid sequence of SEQ ID NO: 256, HC CDR2 comprising the amino acid sequence of SEQ ID NO: 257, HC CDR3 comprising the amino acid sequence of SEQ ID NO: 258, LC CDR1 comprising the amino acid sequence of SEQ ID NO: 259, LC CDR2 comprising the amino acid sequence of SEQ ID NO: 260, and LC CDR3 comprising the amino acid sequence of SEQ ID NO: 261. In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody comprising at least one antigen-specific binding site that specifically binds to IL-4R includes HC CDR1 comprising the amino acid sequence of SEQ ID NO: 262, HC CDR2 comprising the amino acid sequence of SEQ ID NO: 263, HC CDR3 comprising the amino acid sequence of SEQ ID NO: 264, LC CDR1 comprising the amino acid sequence of SEQ ID NO: 265, LC CDR2 comprising the amino acid sequence of SEQ ID NO: 267, and LC CDR3 comprising the amino acid sequence of SEQ ID NO: 268. In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody, which includes at least one antigen-specific binding site that specifically binds to IL-4R, comprises HC CDR1 containing the amino acid sequence of SEQ ID NO: 269, HC CDR2 containing the amino acid sequence of SEQ ID NO: 270, HC CDR3 containing the amino acid sequence of SEQ ID NO: 271, LC CDR1 containing the amino acid sequence of SEQ ID NO: 272, LC CDR2 containing the amino acid sequence of SAS, and LC CDR3 containing the amino acid sequence of SEQ ID NO: 274. In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody, which includes at least one antigen-specific binding site that specifically binds to IL-4R, comprises VH containing the amino acid sequence of SEQ ID NO: 254, and VL containing the amino acid sequence of SEQ ID NO: 255.

[0163] It is known that bispecific antibodies may not retain the same binding properties to each antigen compared to the parent antibody from which they are derived, due to changes in size, titer, structure, arrangement, valency, mobility, geometric shape, and heavy / light chain pairing (see, for example, Brinkmanna et al., The making of bispecific antibodies, MAbs. 2017 Feb-Mar;9(2):182-212).

[0164] However, in some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody maintains a similar range of binding affinity to each antigen (e.g., KLK5 / KLK7 and IL-4) compared to the binding affinity of the antibody from which each of the antigen-specific binding sites of the bispecific antibody originates, corresponds to, or is identical to (also called the "parent antibody of the bispecific antibody") (e.g., difference of 10% or less, difference of 20% or less, difference of 30% or less, difference of 40% or less, difference of 50% or less, difference of 10% or less, difference of less than 10x, difference of less than 9x, difference of less than 8x, difference of less than 7x, difference of less than 6x, difference of less than 5x, difference of less than 4x, difference of less than 3x, difference of less than 2x, or difference of less than 1x). Therefore, in some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody maintains a similar range of binding affinity to KLK5 / KLK7 (e.g., difference of 10% or less, difference of 20% or less, difference of 30% or less, difference of 40% or less, difference of 50% or less, difference of 10% or less, difference of less than 10 times, difference of less than 9 times, difference of less than 8 times, difference of less than 7 times, difference of less than 6 times, difference of less than 5 times, difference of less than 4 times, difference of less than 3 times, difference of less than 2 times, or difference of less than 1 time) compared to the binding affinity of the anti-KLK5 / KLK7 antibody from which the bispecific antibody is derived, corresponding to, or identical to. Therefore, in some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody maintains a similar range of binding affinity to IL-4 or IL-4R (e.g., difference of 10% or less, difference of 20% or less, difference of 30% or less, difference of 40% or less, difference of 50% or less, difference of 10% or less, difference of less than 10x, difference of less than 9x, difference of less than 8x, difference of less than 7x, difference of less than 6x, difference of less than 5x, difference of less than 4x, difference of less than 3x, difference of less than 2x, or difference of less than 1x) compared to the binding affinity of the anti-IL-4 or anti-IL-4R antibody from which the bispecific antibody is derived, corresponds to, or is the same as.

[0165] In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody comprising CDR and / or VH / VL derived from, corresponding to, or identical to, the CDR and / or VH / VL of KLK5 / KLK7-dual-Ab4, and CDR and / or VH / VL derived from, corresponding to, or identical to, the CDR and / or VH / VL of an anti-IL-4R antibody (such as dupilumab), is KL Compared to the binding affinity of dupilumab to KLK5 / KLK7-dual-Ab4 and IL-4R, it maintains a similar range of binding affinity (e.g., difference of ≤10%, ≤20%, ≤30%, ≤40%, ≤50%, ≤10%, ≤10x, ≤9x, ≤8x, ≤7x, ≤6x, ≤5x, ≤4x, ≤3x, ≤2x, or ≤1x). Therefore, in some embodiments, anti-KLK5 / KLK7 includes CDR and / or VH / VL derived from, corresponding to, or identical to, CDR and / or VH / VL of KLK5 / KLK7-dual-Ab4, as well as CDR and / or VH / VL derived from, corresponding to, or identical to, CDR and / or VH / VL of anti-IL-4 or anti-IL-4R antibodies such as dupilumab. +IL-4 bispecific antibody maintains a similar range of binding affinity to KLK5 / KLK7 compared to KLK5 / KLK7-dual-Ab4 (e.g., difference of less than 10%, less than 20%, less than 30%, less than 40%, less than 50%, less than 10%, less than 10x, less than 9x, less than 8x, less than 7x, less than 6x, less than 5x, less than 4x, less than 3x, less than 2x, or less than 1x).Therefore, in some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody comprising CDR and / or VH / VL derived from, corresponding to, or identical to, the CDR and / or VH / VL of KLK5 / KLK7-dual-Ab4, and CDR and / or VH / VL derived from, corresponding to, or identical to, the CDR and / or VH / VL of an anti-IL-4 or anti-IL4R antibody such as dupilumab, is derived from, or is identical to, the bispecific antibody. It maintains a similar range of binding affinity to IL-4 or IL-4R (e.g., difference of ≤10%, ≤20%, ≤30%, ≤40%, ≤50%, ≤10%, ≤10, ≤9, ≤8, ≤7, ≤6, ≤5, ≤4, ≤3, ≤3, ≤3, ≤3, ≤3, ≤3, ≤4, ≤3, ≤3, ≤3, ≤3, ≤3, ≤3, ≤3, ≤3, ≤3, ≤3, or ≤3,

[0166] In some embodiments, the anti-KLK5 / KLK7+IL-4 bispecific antibody retains a similar range of inhibitory activity against each antigen (e.g., KLK5 / KLK7 and IL-4) compared to the inhibitory activity of an antibody from which each of the antigen-specific binding sites of the bispecific antibody originates, corresponds to, or is identical to (e.g., difference of ≤10%, ≤20%, ≤30%, ≤40%, ≤50%, ≤10%, ≤10, ≤9, ≤8, ≤7, ≤6, ≤5, ≤4, ≤3 In some embodiments, an anti-KLK5 / KLK7+IL-4 bispecific antibody comprising a CDR and / or VH / VL derived from, corresponding to, or identical to, the CDR and / or VH / VL of KLK5 / KLK7-dual-Ab4, and a CDR and / or VH / VL derived from, corresponding to, or identical to, the CDR and / or VH / VL of an anti-IL-4 or anti-IL-4R antibody such as dupilumab, is KLK5 Compared to the inhibitory activity of dupilumab against KLK5 / KLK7 protease and IL-4 signaling, the bispecific antibody and parent antibody retain a similar range of inhibitory activity (e.g., difference of less than 10%, less than 20%, less than 30%, less than 40%, less than 50%, less than 10%, less than 10x, less than 9x, less than 8x, less than 7x, less than 6x, less than 5x, less than 4x, less than 3x, less than 2x, or less than 1x). The inhibitory activity of the bispecific antibody and parent antibody can be measured by any suitable method, e.g., KLK5 / KLK7 protease activity against BOC-Val-Pro-Arg-AMC and / or KHLF-AMC, RT-PCR, Western blotting, etc.

[0167] In some embodiments, one or more anti-KLK5 / KLK7 antibodies may be combined with any other suitable anti-KLK7 antibodies to produce multispecific or bispecific anti-KLK5 / KLK7 antibodies. For example, the anti-KLK5 / KLK7 antibodies described herein (Tables 1a and 1b) can be combined with any other suitable anti-KLK7 antibodies to produce bispecific antibodies. Non-limiting examples of suitable anti-KLK7 antibodies include U.S. Patent Application Publication 2021-0130492, titled "ANTI-KLK7 ANTIBODIES, ANTI-KLK5 ANTIBODIES, MULTISPECIFIC ANTI-KLK5 / KLK7 ANTIBODIES, AND METHODS OF USE," published May 6, 2021; International Patent Application Publication WO2021226695, titled "RECOMBINANT HUMAN ANTIBODIES FOR INHIBITING HUMAN TISSUE KALLIKREIN7 (KLK7) AND USE IN DISEASES RELATED TO THE PROCESS OF SKIN DESQUAMATION," published November 18, 2021; and International Patent Application Publication WO2005075667, titled "DIAGNOSTICS AND THERAPEUTICS FOR DISEASES Associated with Kallikrein7 (KLK7), released on August 18, 2005, the contents of which are incorporated herein by reference.

[0168] In some embodiments, one or more anti-KLK5 / KLK7 antibodies may be combined with any suitable anti-KLK5 antibody to produce multispecific or bispecific anti-KLK5 / KLK7 antibodies. For example, the KLK5 / KLK7 antibodies described herein (Tables 1a and 1b) can be combined with any suitable anti-KLK5 antibody. Non-limiting examples of anti-KLK5 antibodies include: U.S. Patent No. 11,292,828, titled "KLK5 Inhibitory Peptide," filed April 5, 2022; U.S. Patent Publication No. 2022-0306725, titled "KLK5 Inhibitory Peptide," published September 29, 2022; U.S. Patent Publication No. 2019-0078160, titled "Use of KLK5 Antagonists for Treatment of a Disease," published March 14, 2019; International Patent Publication No. WO2021156171, titled "Antibodies Against KLK5," published August 12, 2021; International Patent Publication No. WO2021156170, titled "Antibodies Against KLK5" The information is provided in U.S. Patent Application Publication No. 2021-0301032, titled "ANTI-KLK5 ANTIBODIES AND METHODS OF USE", published on August 12, 2021; U.S. Patent Application Publication No. 2021-0130492, titled "ANTI-KLK7 ANTIBODIES, ANTI-KLK5 ANTIBODIES, MULTISPECIFIC ANTI-KLK5 / KLK7 ANTIBODIES, AND METHODS OF USE", published on May 6, 2021, and the contents of these are incorporated herein by reference.

[0169] In some embodiments, the multispecific antibody comprises 3, 4, 5, 6, 7, 8, or more distinct antigen-specific binding sites. In some embodiments, each distinct antigen-specific binding site of the multispecific antibody targets a different antigen. In some embodiments, each distinct antigen-specific binding site of the multispecific antibody targets a different region of the same antigen. In some embodiments, the multispecific antibody comprises distinct antigen-specific binding sites targeting different antigens and / or distinct antigen-specific binding sites targeting different regions of the same antigen. In some embodiments, the multispecific antibody comprises at least one antigen-specific binding site targeting a first antigen and at least one antigen-specific binding site targeting a second antigen. In some embodiments, the multispecific antibody comprises two or more antigen-specific binding sites targeting different regions of the first antigen and / or two or more antigen-specific binding sites targeting different regions of the second antigen.

[0170] In some embodiments, the multispecific antibody targets two antigens and contains one antigen-specific binding site (1+1) for each antigen. In some embodiments, the multispecific antibody targets two antigens and contains two antigen-specific binding sites (2+2) for each antigen. In some embodiments, the multispecific antibody targets two antigens and contains one antigen-specific binding site for one antigen and two antigen-specific binding sites (1+2) for the other antigen. In some embodiments, the multispecific antibody targets two antigens and contains two antigen-specific binding sites for one antigen and three antigen-specific binding sites (2+3) for the other antigen. In some embodiments, the multispecific antibody targets two antigens and contains three antigen-specific binding sites for one antigen and three antigen-specific binding sites (3+3) for the other antigen.

[0171] In some embodiments, multispecific antibodies lack Fc-mediated effector functions, such as antibody-dependent cell-mediated cytotoxicity (ADCC), antibody-dependent phagocytosis (ADCP), complement fixation, and FcRn-mediated recycling. However, in some embodiments, multispecific antibodies include one or more Fc regions that support Fc-mediated effector functions, such as antibody-dependent cell-mediated cytotoxicity (ADCC), antibody-dependent phagocytosis (ADCP), complement fixation, and FcRn-mediated recycling.

[0172] In some embodiments, the antibodies provided herein are bispecific antibodies. In some embodiments, the bispecific antibody comprises at least two different Fv regions. In some embodiments, the bispecific antibody comprises two different heavy chains and two different light chains. In some embodiments, the bispecific antibody comprises one or more IgG molecules. In some embodiments, the bispecific antibody comprises one or more IgG molecules containing additional antigen-specific binding sites, for example, IgG molecules containing an added or modified Ig-like structure.

[0173] In some embodiments, the bispecific antibody comprises two single-strand variable fragments (scFv) linked via a linker. In some embodiments, the bispecific antibody comprises two single-domain antibodies, such as VH or VL domains, VHH, VNAR, or nanobodies, linked via a linker (e.g., a mobile glycine-rich linker such as a (G4S)3 linker). In some embodiments, the bispecific antibody is in diabodies format, as described, for example, in P. Holliger, T. Prospero, and G. Winter, “Diabodies”: small bivalent and bispecific antibody fragments, Proc Natl Acad Sci US A. 1993 Jul 15;90(14):6444-6448, the full contents of which are incorporated herein by reference in their entirety. In some embodiments, the bispecific antibody is a Fab fusion protein, such as a Fab-Fab fusion protein, a Fab-scFv fusion protein, or a Fab-Fv fusion protein. In some embodiments, the bispecific antibody includes an antigen-binding site, e.g., scFv, which is modified to contain a second distinct antigen-specific binding site, e.g., scFv, as an integral part of the antibody.

[0174] In some embodiments, the bispecific antibody is in a fragment-based format, a symmetric format, or an asymmetric format. In some embodiments, the fragment-based bispecific antibody does not contain an Fc region. In some embodiments, the bispecific antibody is in a tandem VHH, tandem scFv, DART, diabody, F(ab)2, scFv-Fab, tandem VHH, (scFv)2-Fab, or tandem diabody format. In some embodiments, the bispecific antibody is in an asymmetric format selected from rat-mouse hybrid IgG, hetero-H HL exchange and / or assembled IgG, hetero-H forced HL IgG, cH IgG, hetero-H CrossMab, scFv-Fab IgG, DART-Fc, LP-DART, CODV-Fab-TL, HLE-BiTE, and F(ab)3CrossMab format. In some embodiments, the bispecific antibodies are symmetrical formats selected from IgG-(scFv)2, Bs4Ab, DVD-Ig, tetravalent DART-Fc, (scFV)4-Fc, CODV-Ig, two-in-one, mAb2, F(ab)4 CrossMab, and tandem VHH-Fc formats.

[0175] In some embodiments, bispecific antibodies are manipulated to facilitate formation via knobs-into-holes techniques, for example, to promote heterodimerization. Using knobs-into-holes technology, in some embodiments, bispecific IgG molecules, trivalent Ig-like antibodies, bispecific Fc and CH3 fusion proteins, and other formats may be generated, as described in Ridgway JB, et al., 'Knobs-into-holes' engineering of antibody CH3 domains for heavy chain heterodimerization. Protein Eng 1996;9:617-21; Atwell S, et al., Stable heterodimers from remodeling the domain interface of a homodimer using a phage display library, J Mol Biol 1997;270:26-35; and Merchant AM, et al., An efficient route to human bispecific IgG, Nat Biotechnol 1998;16:677-681, the entire contents of each of these are incorporated herein by reference.

[0176] In some embodiments, bispecific antibodies lack Fc-mediated effector functions, such as antibody-dependent cell-mediated cytotoxicity (ADCC), antibody-dependent phagocytosis (ADCP), complement fixation, and / or FcRn-mediated recycling. However, in some embodiments, bispecific antibodies include one or more Fc regions that support Fc-mediated effector functions, such as antibody-dependent cell-mediated cytotoxicity (ADCC), antibody-dependent phagocytosis (ADCP), complement fixation, and FcRn-mediated recycling.

[0177] III. Antibody Preparation The antibodies described herein can be prepared by any method known in the art. For example, see Harlow and Lane, (1998) Antibodies: A Laboratory Manual, Cold Spring Harbor Laboratory, New York.

[0178] In some embodiments, antibodies specific to a target antigen (e.g., KLK5 and / or KLK7) can be produced by conventional hybridoma techniques. Using a full-length target antigen or a fragment thereof, optionally coupled to a carrier protein such as KLH, a host animal can be immunized to produce antibodies that bind to that antigen. The routes and schedules of immunization of the host animal are generally consistent with established conventional techniques for antibody stimulation and production, as further described herein. Common techniques for mouse, humanization, and human antibody production are known in the art and are described herein. Any mammalian subject, including human or human-derived antibody-producing cells, is intended to be manipulated to serve as a basis for the production of mammalian, e.g., human hybridoma cell lines. Typically, the host animal is inoculated intraperitoneally, intramuscularly, orally, subcutaneously, plantarly, and / or intradermally with a certain amount of immunogen, including those described herein.

[0179] In some embodiments, bispecific antibodies targeting two different antigens (e.g., KLK5 / KLK7 and IL-4 or their receptors) can be prepared by preferred methods, such as those described in, for example, Nature Reviews Drug Discovery volume 18, pages 585-608 (2019) and Brinkmann U and Kontermann EE, The making of bispecific antibodies, MAbs. 2017 Feb / Mar;9(2):182-212. In some embodiments, anti-KLK5 / KLK7 + IL-4 or its receptor bispecific antibody can be produced by expressing VH and / or VL of one arm and VH and / or VL of the other arm in one or more host cells. In some embodiments, the coding sequence of the arm is described herein to specifically bind to KLK5 / KLK7. In some embodiments, the coding sequence of the arm that specifically binds to IL-4 or its receptor can be developed to express the amino acid sequence of the IL-4 targeted antibody described herein. In some embodiments, different chains of bispecific antibodies can be assembled to form a bispecific antibody using any preferred method.

[0180] If desired, the antibody of interest (monoclonal or polyclonal) (e.g., produced by a hybridoma) may be sequenced, and the polynucleotide sequence may then be cloned into a vector for expression or proliferation. The sequence encoding the antibody of interest may be maintained in a vector within host cells, and then the host cells may be grown and frozen for future use. Alternatively, the polynucleotide sequence may be used for genetic engineering to "humanize" the antibody, or to improve the antibody's affinity (affinity maturation) or other properties. For example, when the antibody is to be used in clinical trials and human treatments, the constant region may be manipulated to more closely resemble the human constant region in order to avoid an immune response. Genetic engineering of the antibody sequence may be desirable to obtain higher affinity and higher efficacy against the target antigen. It will be apparent to those skilled in the art that one or more polynucleotide changes can be made to an antibody while still maintaining its binding specificity to the target antigen.

[0181] In other embodiments, fully human antibodies can be obtained by using commercially available mice engineered to express specific human immunoglobulin proteins. Transgenic animals designed to produce a more desirable (e.g., fully human antibodies) or more robust immune response can also be used to produce humanized antibodies or human antibodies. Examples of such techniques include Xenomouse® from Amgen, Inc. (Fremont, CA), HuMAb-Mouse® and TC mice® from Medarex, Inc. (Princeton, NJ), or H2L2 mice from Harbor Antibodies BV (Holland). Alternatively, antibodies can be recombinantly produced by phage display or yeast technology. See, for example, U.S. Patents 5,565,332, 5,580,717, 5,733,743, and 6,265,150, and Winter et al., (1994) Annu. Rev. Immunol. 12:433-455. Alternatively, phage display technology (McCafferty et al., (1990) Nature 348:552-553) can be used to generate human antibodies and antibody fragments in vitro from a gene repertoire of immunoglobulin variable (V) domains derived from non-immunized donors.

[0182] Antigen-binding fragments of intact antibodies (full-length antibodies) can be prepared by conventional methods. For example, the F(ab')2 fragment can be produced by pepsin digestion of the antibody molecule, and the Fab fragment can be produced by reducing the disulfide crosslinks of the F(ab')2 fragment. Genetically modified antibodies, such as humanized antibodies, chimeric antibodies, single-chain antibodies, and bispecific antibodies, can be produced, for example, by conventional recombination techniques. In one example, DNA encoding a monoclonal antibody specific to a target antigen can be readily isolated and sequenced using conventional procedures (for example, by using oligonucleotide probes that can specifically bind to the genes encoding the heavy and light chains of the monoclonal antibody). Hybridoma cells serve as a preferred source of such DNA. After isolation, the DNA may be placed in one or more expression vectors, which are then transfected into host cells that do not otherwise produce immunoglobulin proteins, such as E. coli cells, Simian COS cells, Chinese hamster ovary (CHO) cells, human HEK293 cells, or myeloma cells, to synthesize monoclonal antibodies in recombinant host cells. See, for example, PCT Publication WO87 / 04462. The DNA can then be modified, for example, by substituting the coding sequences of the constant domains of human heavy and light chains for homologous mouse sequences (Morrison et al., (1984) Proc. Nat. Acad. Sci. 81:6851), or by covalently bonding all or part of the coding sequence of a non-immunoglobulin polypeptide to the immunoglobulin coding sequence. In this way, genetically modified antibodies, such as “chimeric” or “hybrid” antibodies, which have binding specificity to target antigens, can be prepared.

[0183] Single-chain antibodies can be prepared via recombinant technology by linking nucleotide sequences encoding a heavy chain variable region and a light chain variable region. Preferably, a movable linker is incorporated between the two variable regions.

[0184] Antibodies known in the art and obtained according to methods described herein can be characterized using methods well known in the art. For example, one method is to identify the epitopes to which the antigen binds, or "epitope mapping." There are many methods known in the art for mapping and characterizing the location of epitopes on proteins, including analysis of the crystalline structure of antibody-antigen complexes, competitive assays, gene fragment expression assays, and synthetic peptide-based assays, for example, Chapter 11 of Harlow and Lane, Using Antibodies, a Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, 1999. In one example, epitope mapping can be achieved using H / D-Ex (hydrogen-deuterium exchange) combined with proteolysis and mass spectrometry. In an additional example, epitope mapping can be used to determine the sequences to which the antibody binds. Epitopes can be linear epitopes, i.e., epitopes contained in a sequence of amino acids, or stereostructural epitopes formed by three-dimensional interactions of amino acids that are not necessarily contained in a sequence (linear sequence of primary structure). Peptides of various lengths (e.g., at least 4-6 amino acid lengths) can be isolated or synthesized (e.g., by recombination) and used in antibody binding assays. In another example, the epitope to which the antibody binds can be determined in systematic screening by using duplicate peptides derived from the target antigen sequence and determining antibody binding. In gene fragment expression assays, an open reading frame encoding the target antigen is fragmented either randomly or by a specific gene construction, and the reactivity of the expressed antigen fragment with the antibody being tested is determined. The gene fragment can be generated, for example, by PCR, then transcribed in vitro in the presence of radioactive amino acids and translated into a protein. The binding of the antibody to the radioactively labeled antigen fragment is then determined by immunoprecipitation and gel electrophoresis.Certain epitopes can also be identified by using a large library of random peptide sequences presented on the surface of phage particles (phage libraries). Alternatively, a defined library of overlapping peptide fragments can be tested for binding to a test antibody in a simple binding assay. In further examples, antigen-binding domain mutagenesis, domain swapping experiments, and alanine scanning mutagenesis can be performed to identify the sufficient and / or required residues for epitope binding. Alternatively, a competitive assay can be performed using other antibodies known to bind to the same antigen to determine whether an antibody binds to the same epitope as other antibodies. Competitive assays are well known to those skilled in the art.

[0185] In some cases, the antibodies described herein are prepared by recombinant techniques as illustrated below. The nucleic acids encoding the heavy and light chains of the antibodies described herein can be cloned into a single expression vector, with each nucleotide sequence operably ligated to a suitable promoter. In one example, each of the nucleotide sequences encoding the heavy and light chains is operably ligated to a separate promoter. Alternatively, the nucleotide sequences encoding the heavy and light chains can be operably ligated to a single promoter so that both the heavy and light chains are expressed from the same promoter. If necessary, an intra-sequence ribosome entry site (IRES) can be inserted between the heavy chain coding sequence and the light chain coding sequence.

[0186] In some cases, the nucleotide sequences encoding the two chains of an antibody can be cloned into two vectors, and these can be introduced into the same or different cells. If these two chains are expressed in different cells, each of them can be isolated from the host cell expressing it, and the isolated heavy and light chains can be mixed and incubated under suitable conditions to enable antibody formation.

[0187] Generally, nucleic acid sequences encoding one or all of an antibody chain can be cloned into a suitable expression vector, operably ligated to a suitable promoter, using methods known in the art. For example, the nucleotide sequence and vector can be contacted with restriction enzymes under suitable conditions to create complementary ends on each molecule that can pair with each other and be ligated together. Alternatively, synthetic nucleic acid linkers can be ligated to the ends of the gene. These synthetic linkers contain nucleic acid sequences corresponding to specific restriction sites in the vector. The choice of expression vector / promoter will depend on the type of host cell to be used for antibody production.

[0188] Various promoters can be used for the expression of the antibodies described herein, but are not limited to, the initial promoter of cytomegalovirus (CMV), viral LTRs such as Rous sarcoma virus LTR, HIV-LTR, HTLV-1 LTR, Simian virus 40 (SV40) initial promoter, E. coli lac UV promoter, and herpes simplex virus tk promoter.

[0189] Controllable promoters can also be used. Such controllable promoters include those that use the E. coli-derived lac repressor as a transcription regulator to control transcription from lac operator-containing mammalian cell promoters [[Brown, M. et al., Cell, 49:603-612 (1987)]] and those that use the tetracycline repressor (tetR) [[Gossen, M., and Bujard, H., Proc. Natl. Acad. Sci. USA 89:5547-555115 (1992), Yao, F. et al., Human Gene Therapy, 9:1939-1950 (1998), Shockelt, P., et al., Proc. Natl. Acad. Sci. USA, 92:6522-6526 (1995)]]. Other systems include FK506 dimers, VP16 or p65 using estradiol, RU486, diphenol murislerone, or rapamycin. Among these, derivable systems are available from Invitrogen, Clontech, and Ariad.

[0190] A controllable promoter containing a repressor can be used with the operon. In one embodiment, the lac repressor derived from E. coli can function as a transcription regulator that controls transcription from mammalian cell promoters containing the lac operator [[M. Brown et al., Cell, 49:603-612 (1987)]]; Gossen and Bujard (1992); [[M. Gossen et al., Natl. Acad. Sci. USA, 89:5547-5551 (1992)]]. Combining the tetracycline repressor (tetR) with the transcription activator (VP16) to create the tetR-mammalian cell transcription activator fusion protein, tTa (tetR-VP16), and combining it with the tetO-containing minimal promoter derived from the human cytomegalovirus (hCMV) promoter, a tetR-tet operator system was created for controlling gene expression in mammalian cells. In one embodiment, a tetracycline-inducible switch is used. Tetracycline repressor (tetR) alone can function as a potent trans-regulator for controlling gene expression in mammalian cells, provided that the tetracycline operator is properly positioned downstream of the TATA element of the CMVIE promoter, more so than tetR-mammalian cell transcription factor fusion derivatives (Yao et al., Human Gene Therapy). One particular advantage of this tetracycline-inducible switch is that it does not require the use of tetracycline repressor-mammalian cell transcription activators or repressor fusion proteins, which may in some cases be toxic to cells, to achieve its controllable effect (Gossen 5 et al., Natl. Acad. Sci. USA, 89:5547-5551 (1992); Shockett et al., Proc. Natl. Acad. Sci. USA, 92:6522-6526 (1995)).

[0191] Furthermore, the vector may contain, for example, some or all of the following: selection marker genes, e.g., the neomycin gene for selection of stable or transient transtransitors in mammalian cells; enhancer / promoter sequences from the pre-initial gene of human CMV for high levels of transcription; transcription termination and RNA processing signals from SV40 for mRNA stability; SV40 polyomatous replication origin and ColE1 for proper episomal replication; intrasequence ribosome binding sites (IRESs), versatile multiplexing sites; and T7 and SP6 RNA promoters for in vitro transcription of sense and antisense RNA. Appropriate vectors and methods for generating vectors containing transgenes are well known and available in the art. Examples of polyadenylation signals useful for carrying out the methods described herein include, but are not limited to, the human collagen I polyadenylation signal, the human collagen II polyadenylation signal, and the SV40 polyadenylation signal.

[0192] One or more vectors (e.g., expression vectors) containing nucleic acids encoding any of the antibodies (e.g., nucleic acid coding sequences listed in Table 3) can be introduced into suitable host cells for antibody production. Non-limiting examples of host cells include Chinese hamster ovary (CHO) cells, dhfr-CHO cells, human embryonic kidney (HEK)-293 cells, verda reno (VERO) cells, non-secretory null (NS0) cells, human embryonic retina (PER.C6) cells, Sp2 / 0 cells, baby hamster kidney (BHK) cells, Madin-Darby canine kidney (MDCK) cells, Madin-Darby bovine kidney (MDBK) cells, and SV40-transformed monkey kidney CV1 (COS) cells. In some embodiments, the host cells expressing the antibodies described herein are CHO cells. The host cells can be cultured under conditions suitable for the expression of the antibody or any polypeptide chain thereof. Such antibodies or their polypeptide chains can be recovered by cultured cells (e.g., from cells or culture supernatant) via conventional methods, such as affinity purification. If necessary, the polypeptide chains of the antibodies can be incubated under suitable conditions for a suitable period of time to allow antibody production. In some embodiments, the host cells contain nucleic acids encoding the heavy chain of the antibodies described herein. In some embodiments, the host cells contain nucleic acids encoding the light chain of the antibodies described herein. In some embodiments, the host cells contain nucleic acids encoding both the heavy chain and the light chain.

[0193] In some embodiments, the method for preparing the antibodies described herein involves a recombinant expression vector encoding both the heavy and light chains of the antibodies described herein, as also described herein. The recombinant expression vector can be introduced into suitable host cells (e.g., dhfr-CHO cells) by conventional methods, such as calcium phosphate-mediated transfection. Positive transformant host cells can be selected and cultured under suitable conditions to enable the expression of the two polypeptide chains that form the antibody, and these can be recovered from the cells or culture medium. If necessary, the two chains recovered from the host cells can be incubated under suitable conditions to enable antibody formation.

[0194] In one example, two recombinant expression vectors are provided, one encoding the heavy chain of an antibody and the other encoding the light chain of an antibody. Both recombinant expression vectors can be introduced into suitable host cells (e.g., dhfr-CHO cells) by conventional methods, such as calcium phosphate-mediated transfection.

[0195] Alternatively, each expression vector can be introduced into a suitable host cell. Positive transformants can be selected and cultured under suitable conditions to enable the expression of the antibody polypeptide chain. If two expression vectors are introduced into the same host cell, the antibody produced in the cell can be recovered from the host cell or culture medium. If necessary, the polypeptide chain can be recovered from the host cell or culture medium and then incubated under suitable conditions to enable antibody formation. If two expression vectors are introduced into different host cells, each of them can be recovered from the corresponding host cell or corresponding culture medium. The two polypeptide chains can then be incubated under suitable conditions for antibody formation.

[0196] Using standard molecular biology techniques, recombinant expression vectors are prepared, host cells are transfected, transformants are selected, host cells are cultured, and antibodies are recovered from the culture medium. Some antibodies can be isolated, for example, by affinity chromatography using a protein A or protein G binding matrix.

[0197] Any nucleic acids encoding the heavy chain, light chain, or both of the antibodies described herein (e.g., those provided in Table 3), vectors containing them (e.g., expression vectors), and host cells containing the vectors are all within the scope of this disclosure. [Table 3-1] [Table 3-2]

[0198] In some embodiments, the Disclosure provides isolated nucleic acids comprising a sequence that is at least 60% (e.g., 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identical to any one of SEQ ID NOs. In some embodiments, the Disclosure provides isolated nucleic acids comprising a sequence that is at least 60% (e.g., 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identical to any one of SEQ ID NOs. In some embodiments, the Disclosure provides isolated nucleic acids comprising sequences that are at least 60% (e.g., 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identical to any one of sequence numbers 22-27 or 275-276.

[0199] In some embodiments, this disclosure provides an expression vector encoding an anti-KLK5 / KLK7 antibody as described herein. In some embodiments, the expression vector comprises an isolated nucleic acid that is at least 60% (e.g., 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identical to any one of SEQ ID NOs. In some embodiments, the expression vector comprises an isolated nucleic acid that is at least 60% (e.g., 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identical to any one of SEQ ID NOs. In some embodiments, the expression vector comprises an isolated nucleic acid that is at least 60% (e.g., 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identical to any one of sequence numbers 22-27 or 275-276.

[0200] In some embodiments, the anti-KLK5 / KLK7 antibodies described herein are produced by expressing in recombinant cells (i) an isolated nucleic acid that is at least 60% (e.g., 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identical to any one of SEQ ID NOs. 22, 24, 26, or 27, and / or (ii) an isolated nucleic acid that is at least 60% (e.g., 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identical to any one of SEQ ID NOs. 23, 25, 275, or 276.

[0201] In some embodiments, the anti-KLK5 / KLK7 antibodies described herein are produced by expressing in recombinant cells an expression vector comprising (i) an isolated nucleic acid that is at least 60% (e.g., 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identical to any one of SEQ ID NOs. 22, 24, 26, and 27, and / or (ii) an isolated nucleic acid that is at least 60% (e.g., 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) identical to any one of SEQ ID NOs. 23, 25, 275, or 276.

[0202] In some embodiments, the antibodies described herein are used to modulate the activity or function of at least one gene, protein, and / or nucleic acid. In some embodiments, the molecular payload is responsible for the modulation of the gene, protein, and / or nucleic acid. The molecular payload may be a small molecule, protein, nucleic acid, oligonucleotide, or any molecular entity capable of modulating the activity or function of an intracellular gene, protein, and / or nucleic acid.

[0203] In some embodiments, the multispecific antibody includes the direct fusion or linkage of different antigen-specific binding sites. In some embodiments, the multispecific antibody includes an immunoglobulin-derived heteromerized domain for generating the multispecific antibody. In some embodiments, the multispecific antibody can be formed by the co-expression of different heavy chains and two different light chains. In some embodiments, the multispecific antibody can be formed by the co-expression of different heavy chains and a common light chain. In some embodiments, the multispecific antibody includes, for example, an engineered CH1 domain (the first constant Ig domain of the heavy chain) that promotes appropriate heavy-light chain pairing in such co-expression systems, as disclosed, for example, in International Patent Application Publication WO2021067404, “CH1 DOMAIN VARIANTS ENGINEERED FOR PREFERENTIAL LIGHT CHAIN ​​PAIRING AND MULTISPECIFIC ANTIBODIES COMPRISING THE SAME,” published April 8, 2021, which is incorporated herein by reference.

[0204] In some embodiments, the multispecific antibody includes a variant CH1 domain that pairs (e.g., preferentially pairs) with a particular variant CL domain. For example, in some embodiments, the multispecific antibody includes a heavy chain containing a variant CH1 domain that preferentially pairs with a variant CLK or CLλ domain. Non-limiting examples of such multispecific antibody configurations are provided in International Patent Application Publication WO2022150787, “VARIANT CH1 DOMAINS AND VARIANT CL DOMAINS ENGINEERED FOR PREFERENTIAL CHAIN ​​PAIRING AND MULTI-SPECIFIC ANTIBODIES COMPRISING THE SAME,” published July 14, 2022, which is incorporated herein by reference in its entirety.

[0205] In some embodiments, multispecific antibodies include a variant CH3 domain that preferentially forms CH3-CH3 heterodimers over CH3-CH3 homodimers. The incorporation of such a variant CH3 domain promotes, for example, heterodimerization of different antibodies to form multispecific antibodies. Non-limiting examples of such multispecific antibody configurations are provided in International Patent Application Publication WO2022150785, “VARIANT CH3 DOMAINS ENGINEERED FOR PREFERENTIAL CH3 HETERODIMERIZATION, MULTI-SPECIFIC ANTIBODIES COMPRISING THE SAME, AND METHODS OF MAKING THEREOF,” published July 14, 2022, the entire content of which is incorporated herein by reference.

[0206] In some embodiments, bispecific antibodies involve the direct fusion or linkage of different antigen-specific binding sites. In some embodiments, bispecific antibodies include an immunoglobulin-derived heterodimerizing domain for generating bispecific antibodies. For example, in some embodiments, bispecific antibodies can be formed by the co-expression of two different heavy chains and two different light chains. In some embodiments, bispecific antibodies can be formed by the co-expression of different heavy chains and a common light chain. In some embodiments, the fusion of two antibody-producing cell lines allows for the combination of heavy and light chains of two different antibodies, resulting in a bispecific antibody containing the heavy and light chains of a first antibody and the heavy and light chains of a second antibody. In some embodiments, the heavy chain constant region and the light chain constant region are of the same isotype. In some embodiments, the heavy chain constant region and the light chain constant region are of different isotypes.

[0207] In some embodiments, the bispecific antibody includes variant heavy and / or light chains that force correct assembly between the two heavy chains and the homologous heavy and light chains, or facilitate the purification of the correctly assembled bispecific antibody (see, for example, Figures 3 and 4 and Table 1 in Brinkmann U and Kontermann EE, The making of bispecific antibodies, MAbs. 2017 Feb / Mar;9(2):182-212). In some embodiments, the bispecific antibody is formed using, for example, a knobs-into-holes technique to facilitate heterodimerization. Using knobs-into-holes technology, in some embodiments, bispecific IgG molecules, trivalent Ig-like antibodies, bispecific Fc and CH3 fusion proteins, and other formats may be generated, as described in Ridgway JB, et al., 'Knobs-into-holes' engineering of antibody CH3 domains for heavy chain heterodimerization. Protein Eng 1996;9:617-21; Atwell S, et al., Stable heterodimers from remodeling the domain interface of a homodimer using a phage display library, J Mol Biol 1997;270:26-35; and Merchant AM, et al., An efficient route to human bispecific IgG, Nat Biotechnol 1998;16:677-681, the entire contents of each of these are incorporated herein by reference.

[0208] In some embodiments, the bispecific antibody includes, for example, an engineered CH1 domain (the first constant Ig domain of the heavy chain) that promotes appropriate heavy-light chain pairing in such co-expression systems, as disclosed, for example, in International Patent Application Publication WO2021067404, "CH1 DOMAIN VARIANTS ENGINEERED FOR PREFERENTIAL LIGHT CHAIN ​​PAIRING AND MULTISPECIFIC ANTIBODIES COMPRISING THE SAME," published April 8, 2021, which is incorporated herein by reference.

[0209] In some embodiments, the bispecific antibody includes a variant CH1 domain that pairs (e.g., preferentially pairs) with a particular variant CL domain. For example, in some embodiments, the bispecific antibody includes a heavy chain containing a variant CH1 domain that preferentially pairs with a variant CLK or CLλ domain. Non-limiting examples of such bispecific antibody configurations are provided in International Patent Application Publication WO2022150787, “VARIANT CH1 DOMAINS AND VARIANT CL DOMAINS ENGINEERED FOR PREFERENTIAL CHAIN ​​PAIRING AND MULTI-SPECIFIC ANTIBODIES COMPRISING THE SAME,” published July 14, 2022, which is incorporated herein by reference in its entirety.

[0210] In some embodiments, bispecific antibodies include a variant CH3 domain that preferentially forms CH3-CH3 heterodimers over CH3-CH3 homodimers. The incorporation of such a variant CH3 domain promotes, for example, heterodimerization of different antibodies to form bispecific antibodies. Non-limiting examples of such bispecific antibody configurations are provided in International Patent Application Publication WO2022150785, “VARIANT CH3 DOMAINS ENGINEERED FOR PREFERENTIAL CH3 HETERODIMERIZATION, MULTI-SPECIFIC ANTIBODIES COMPRISING THE SAME, AND METHODS OF MAKING THEREOF,” published July 14, 2022, the entire content of which is incorporated herein by reference.

[0211] In some embodiments, bispecific antibodies may be formed using non-immunoglobulin heterodimerization modules to combine different antigen-specific binding sites in a non-covalent or covalent manner. For example, in some embodiments, bispecific antibodies are formed via a dock-and-lock method (DNL) that utilizes a heterodimer assembly of a regulatory subunit of cAMP-dependent protein kinase (PKA) and the anchoring domain (AD) of an A kinase anchor protein (AKAP). In some embodiments, bispecific antibodies may be formed using non-immunoglobulin heterodimerization modules such as the Balanase-Bulster system, an adapter / docking tag module based on a mutant RNase I fragment, and a SNARE module based on the interaction of three protein syntaxins, synaptobrevin and SNAP25, to combine different antigen-specific binding sites.

[0212] IV. Pharmaceutical Compositions The antibodies described herein, as well as the encoded nucleic acids or nucleic acid sets, vectors containing them, or host cells containing the vectors, can be mixed with pharmaceutically acceptable carriers (excipients) to form pharmaceutical compositions for use in the treatment of target diseases. “Acceptable” means that the carrier must be compatible with (and preferably capable of stabilizing) the active ingredient of the composition and must not be harmful to the subject being treated. pharmaceutically acceptable excipients (carriers) include buffers, which are well known in the art. See, for example, Remington: The Science and Practice of Pharmacy 20th Ed. (2000), Lippincott Williams and Wilkins, Ed. KE. Hoover.

[0213] An anti-KLK5 / KLK7+IL-4 bispecific antibody containing at least one antigen-specific binding site comprising a pharmaceutical composition disclosed herein may further comprise a suitable buffer. The buffer is a weak acid or base used to maintain the pH of the solution near a selected value after the addition of another acid or base. In some examples, the buffers disclosed herein may be buffers capable of maintaining physiological pH regardless of changes in carbon dioxide concentration (produced by cellular respiration). Exemplary buffers include, but are not limited to, HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid) buffer, Dulbecco's phosphate-buffered saline (DPBS) buffer, or phosphate-buffered saline (PBS) buffer. Such buffers may comprise disodium hydrogen phosphate and sodium chloride, or potassium dihydrogen phosphate and potassium chloride.

[0214] The pharmaceutical compositions described herein comprise one or more suitable salts. Salts are ionic compounds that can be formed by the neutralization reaction of acids and bases. (Skoog, DA; West, DM; Holler, JF; Crouch, SR (2004). “chapters 14-16”. Fundamentals of Analytical Chemistry (8th ed.)). Salts consist of a related number of cations (positively charged ions) and anions (negative ions) such that the product is electrically neutral (has no net charge).

[0215] In some embodiments, the pharmaceutical composition may contain a pharmaceutically acceptable carrier, excipient, or stabilizer in the form of a lyophilized formulation or aqueous solution. (Remington: The Science and Practice of Pharmacy 20th Ed. (2000) Lippincott Williams and Wilkins, Ed. KE. Hoover). In some embodiments, the pharmaceutical composition may be formulated for intravenous injection. In some embodiments, the pharmaceutical composition may be formulated for subcutaneous injection.

[0216] Pharmaceutical compositions used for in vivo administration must be sterile. This can be easily achieved, for example, by filtration through a sterile filtration membrane. Therapeutic antibody compositions are generally placed in containers with sterile access ports, such as intravenous or subcutaneous solution bags or vials with a stopper that can be punctured with a subcutaneous needle.

[0217] V. How to use In certain embodiments, the Disclosure provides methods and related compositions for treating conditions associated with KLK5 and KLK7-related dysregulation, including, for example, Netherton syndrome, atopic dermatitis (with and without filaggrin mutations), eosinophilic esophagitis, prurigo nodosa, chronic pruritus of unknown cause (CPUO), dry skin, asthma (e.g., specifically KLK5-related asthma), ichthyosis vulgaris, and itchy skin. In some embodiments, Th2 cytokines (e.g., IL-13, IL-4, IL-5, IL-6, IL-9, IL-31, and IL-17E (IL-25)) are also involved in the KLK5 and KLK7-related dysregulation conditions described herein. Accordingly, in some embodiments, the Disclosure provides methods and compositions for inhibiting KLK5 and KLK7, as well as IL-4. In some embodiments, this disclosure provides bispecific antibodies for inhibiting KLK5 and KLK7, as well as IL-4, for treating the conditions provided herein.

[0218] Aspects of this disclosure relate to methods and compositions useful for promoting proper barrier function (e.g., epidermal barrier function) (e.g., anti-KLK5 / KLK7 dual inhibitory antibodies or anti-KLK5 / KLK7+IL-4 bispecific antibodies). Overactive kallikrein 5 / 7 is the cause of both genetic and spontaneous disruption of epidermal barrier function and is associated with related disorders such as Netherton syndrome, eosinophilic esophagitis, and atopic dermatitis. Accordingly, in some embodiments, the methods provided herein involve administering an effective amount of one or more anti-KLK5 / KLK7 antibodies or one or more anti-KLK5 / KLK7+IL-4 bispecific antibodies provided herein to a subject for the purpose of restoring the epithelial barrier in a subject that requires such restoration. In other embodiments, methods are provided for addressing one or more aspects of modified barrier function. For example, in some embodiments, the method provided herein involves administering an effective amount of one or more anti-KLK5 / KLK7 antibodies or one or more anti-KLK5 / KLK7+IL-4 bispecific antibodies provided herein to a subject for the purpose of reducing skin infiltration in a subject that requires such reduction. In some embodiments, the method provided herein involves administering an effective amount of one or more anti-KLK5 / KLK7 antibodies or one or more anti-KLK5 / KLK7+IL-4 bispecific antibodies provided herein to a subject for the purpose of reducing epithelial inflammation in a subject that requires such reduction. In some embodiments, the method provided herein involves administering an effective amount of one or more anti-KLK5 / KLK7 antibodies or one or more anti-KLK5 / KLK7+IL-4 bispecific antibodies provided herein to a subject for the purpose of reducing epithelial permeability. In some embodiments, the methods provided herein involve administering an effective amount of the anti-KLK5 / KLK7 antibody or one or more anti-KLK5 / KLK7+IL-4 bispecific antibodies provided herein to a subject for the purpose of reducing parakeratosis in a subject that requires such reduction.In some embodiments, the methods provided herein involve administering an effective amount of one or more anti-KLK5 / KLK7 antibodies or one or more anti-KLK5 / KLK7+IL-4 bispecific antibodies provided herein to a subject for the purpose of reducing inflammatory cytokines in the skin. In some embodiments, the methods provided herein involve administering an effective amount of one or more anti-KLK5 / KLK7 antibodies or one or more anti-KLK5 / KLK7+IL-4 bispecific antibodies provided herein to a subject for the purpose of reducing transepidermal water loss.

[0219] Further aspects of this disclosure relate to methods and compositions useful for treating atopic dermatitis (e.g., anti-KLK5 / KLK7 antibodies or anti-KLK5 / KLK7 + IL-4 bispecific antibodies). Atopic dermatitis (AD), also known as eczema, is a common chronic pruritic inflammatory skin disease. In some embodiments, atopic dermatitis develops in subjects during infancy or early childhood (e.g., 2 years of age or about 2 years of age). Therefore, in some embodiments, the methods provided herein are useful for treating subjects having atopic dermatitis who are 2 years of age or older. Atopic dermatitis may be associated with elevated total serum IgE levels. Therefore, in some embodiments, the methods provided herein are useful for treating subjects having atopic dermatitis whose total serum IgE levels are elevated (e.g., compared to normal IgE levels in subjects without atopic dermatitis or related conditions). In some embodiments, atopic dermatitis is associated with chronic recurrent skin inflammation, impaired epidermal barrier function (e.g., leading to dry skin), and / or IgE-mediated sensitization to allergens such as food and environmental allergens. Accordingly, in some embodiments, methods for treating subjects having atopic dermatitis are provided herein, comprising administering to the subject an effective amount of anti-KLK5 / KLK7 antibody or one or more anti-KLK5 / KLK7+IL-4 bispecific antibodies. Furthermore, in some embodiments, the subject to be treated exhibits chronic recurrent skin inflammation, impaired epidermal barrier function, and / or IgE-mediated sensitization to allergens.

[0220] In some embodiments, administration of the anti-KLK5 / KLK7+IL-4 bispecific antibody described herein to subjects having AD reduces ear thickness in the subjects by more than 10%, more than 20%, more than 30%, more than 40%, more than 50%, more than 60%, more than 70%, more than 80%, more than 90%, or more than 100% compared to subjects receiving one of the parental antibodies of the bispecific antibody (e.g., parental anti-KLK5 / KLK7 antibody, parental anti-IL-4 antibody, or anti-IL4R antibody).

[0221] In some embodiments, administration of the anti-KLK5 / KLK7+IL-4 bispecific antibody described herein to subjects having AD reduces skin erythema / hemorrhage in the subjects by more than 10%, more than 20%, more than 30%, more than 40%, more than 50%, more than 60%, more than 70%, more than 80%, more than 90%, or more than 100% compared to subjects receiving one of the parent antibodies of the bispecific antibody (e.g., parent anti-KLK5 / KLK7 antibody, parent anti-IL-4 antibody, or anti-IL4R antibody).

[0222] In some embodiments, administration of the anti-KLK5 / KLK7+IL-4 bispecific antibody described herein to subjects having AD reduces skin peeling / erosion in the subjects by more than 10%, more than 20%, more than 30%, more than 40%, more than 50%, more than 60%, more than 70%, more than 80%, more than 90%, or more than 100% compared to subjects receiving one of the parent antibodies of the bispecific antibody (e.g., parent anti-KLK5 / KLK7 antibody, parent anti-IL-4 antibody, or anti-IL4R antibody).

[0223] In some embodiments, administration of the anti-KLK5 / KLK7+IL-4 bispecific antibody described herein to subjects having AD reduces skin edema in the subjects by more than 10%, more than 20%, more than 30%, more than 40%, more than 50%, more than 60%, more than 70%, more than 80%, more than 90%, or more than 100% compared to subjects receiving any one of the parent antibodies of the bispecific antibody (e.g., parent anti-KLK5 / KLK7 antibody, parent anti-IL-4 antibody, or anti-IL4R antibody).

[0224] In some embodiments, administration of the anti-KLK5 / KLK7+IL-4 bispecific antibody described herein to subjects having AD reduces skin scaling / dryness in the subjects by more than 10%, more than 20%, more than 30%, more than 40%, more than 50%, more than 60%, more than 70%, more than 80%, more than 90%, or more than 100% compared to subjects receiving any one of the parental antibodies of the bispecific antibody (e.g., parental anti-KLK5 / KLK7 antibody or parental anti-IL-4 antibody or anti-IL4R antibody).

[0225] In some embodiments, administration of the anti-KLK5 / KLK7+IL-4 bispecific antibody described herein to subjects with AD reduces the severity of skin lesions in subjects by more than 10%, more than 20%, more than 30%, more than 40%, more than 50%, more than 60%, more than 70%, more than 80%, more than 90%, or more than 100% compared to subjects receiving one of the parent antibodies of the bispecific antibody (e.g., parent anti-KLK5 / KLK7 antibody or parent anti-IL-4 antibody or anti-IL4R antibody). The severity of skin lesions (four parameters: erythema / hemorrhage, edema, peeling / erosion, and scaling / dryness) is assessed by signs of the ear, neck, and dorsal skin. The overall clinical skin severity score is defined as the sum of the individual scores (0: none, 1: mild, 2: moderate, 3: severe).

[0226] Further aspects of this disclosure relate to methods and compositions useful for treating subjects having Netherton syndrome (e.g., anti-KLK5 / KLK7 antibodies or anti-KLK5 / KLK7+IL-4 bispecific antibodies). In some embodiments, methods for treating subjects having Netherton syndrome are provided herein, comprising administering an effective amount of one or more anti-KLK5 / KLK7 antibodies or one or more anti-KLK5 / KLK7+IL-4 bispecific antibodies to the subject. Netherton syndrome is a rare and severe autosomal recessive skin disorder. In some embodiments, Netherton syndrome is associated with congenital erythroderma, specific hair shaft abnormalities, and / or atopic findings with elevated IgE levels (e.g., compared to normal IgE levels in subjects without Netherton syndrome or related conditions). In some embodiments, subjects with Netherton syndrome exhibit atopic findings including eczematous rash, atopic dermatitis, pruritus, hay fever, angioedema, urticaria, high levels of serum IgE, and / or eosinophilia. In some embodiments, Netherton syndrome is caused by mutations in the serine protease inhibitor Cazal type 5 (SPINK5) gene, which encodes a protease inhibitor lymphoepithelial Cazal type-related inhibitor. In some embodiments, the absence of this protease inhibitor leads to hyperactivity of epidermal proteases, followed by stratum corneum exfoliation. Therefore, in some embodiments, the methods provided herein involve administering an effective amount of one or more anti-KLK5 / KLK7 antibodies or one or more anti-KLK5 / KLK7+IL-4 bispecific antibodies provided herein to a subject for the purpose of improving one or more aspects or symptoms associated with Netherton syndrome (e.g., atopic symptoms, e.g., cutaneous rash, scaling, stratum corneum exfoliation).

[0227] Further embodiments of this disclosure relate to methods and compositions useful for treating eosinophilic esophagitis (e.g., anti-KLK5 / KLK7 antibodies or anti-KLK5 / KLK7 + IL-4 bispecific antibodies). In some embodiments, symptoms of eosinophilic esophagitis include difficulty eating, growth retardation, vomiting, epigastric or chest pain, dysphagia, and food impaction. In some embodiments, subjects with eosinophilic esophagitis are young males with a relatively high predisposition to atopic diseases. In some embodiments, subjects with eosinophilic esophagitis are diagnosed by endoscopic and / or biopsy findings of isolated eosinophils in the esophagus. In some embodiments, eosinophilic esophagitis is histologically defined by the presence of proliferative changes, which in some embodiments include thickening of the basal epithelial layer and / or elongation of the papillae, at least 24 eosinophils per high-magnification field of view of the distal esophagus, and / or absence of eosinophilia in any other intestinal segment evaluated. In some embodiments, subjects with eosinophilic esophagitis have low levels or deficiencies of a specific serine protease inhibitor belonging to the lymphoepithelial Cazal-type inhibitor protein family, such as SPINK7, as observed in esophageal biopsy, for example. In some embodiments, eosinophilic esophagitis is distinguished from reflux esophagitis based on the degree of mucosal eosinophilia and lack of response to acid suppression. In some embodiments, methods for treating subjects with eosinophilic esophagitis are provided herein, comprising administering an effective dose of one or more anti-KLK5 / KLK7 antibodies or one or more anti-KLK5 / KLK7+IL-4 bispecific antibodies to the subject.

[0228] Further aspects of this disclosure relate to methods and compositions useful for treating prurigo nodularis (e.g., anti-KLK5 / KLK7 antibodies or anti-KLK5 / KLK7 + IL-4 bispecific antibodies). Prurigo nodularis is a chronic inflammatory skin disease characterized by symmetrically distributed, extremely itchy rashes, most commonly appearing on the arms, legs, upper back, and / or abdomen. In some embodiments, prurigo nodularis appears alone. However, in some embodiments, prurigo nodularis is associated with other skin diseases or medical conditions, such as cancer, diabetes, chronic kidney disease, or AIDS. In some embodiments, alterations in the function of the immune and nerve systems in the skin are thought to be associated with increased sensation of itchiness (pruritus) leading to frequent scratching behavior; frequent scratching and picking of such skin contributes to further thickening and formation of the lesions. Accordingly, in some embodiments, methods for treating subjects with nodular prurigo are provided herein, which include administering an effective amount of one or more anti-KLK5 / KLK7 antibodies or one or more anti-KLK5 / KLK7+IL-4 bispecific antibodies to the subject.

[0229] Further embodiments of this disclosure relate to methods and compositions useful for treating chronic pruritus (e.g., anti-KLK5 / KLK7 antibodies or anti-KLK5 / KLK7+IL-4 bispecific antibodies). In some embodiments, chronic pruritus is associated with itching lasting longer than six weeks (e.g., up to three months, up to six months, up to one year or longer). In some embodiments, chronic pruritus occurs in association with chronic kidney disease, hepatobiliary disease, and potentially unrelated diseases, including neuropathic entities such as brachioradial pruritus and dysalgesia. In some embodiments, chronic pruritus of unknown etiology (CPUO) is established when the underlying cause of the itching cannot be identified. In some embodiments, chronic pruritus is associated with severe itching and significant scratching lesions. In some embodiments, methods for treating subjects with chronic pruritus, including CPUO, are provided herein, comprising administering an effective amount of one or more anti-KLK5 / KLK7 antibodies or one or more anti-KLK5 / KLK7+IL-4 bispecific antibodies to the subject.

[0230] Further aspects of this disclosure relate to methods and compositions useful for treating ichthyosis vulgaris (e.g., anti-KLK5 / KLK7 antibodies or anti-KLK5 / KLK7 + IL-4 bispecific antibodies). In some embodiments, ichthyosis vulgaris is caused by heterozygous mutations in the filaggrin gene. In some embodiments, subjects with homozygous or compound heterozygous mutations in this gene have a more severe phenotype. In some embodiments, ichthyosis vulgaris is histologically characterized by the absence or reduction of keratohyalin granules in the epidermis and mild keratosis. Keratohyalin contains a histidine-rich protein that is a precursor form (profilaggrin) of filaggrin, which is a keratin filament aggregate protein. In some embodiments, profilaggrin and filaggrin are reduced or absent in subjects having ichthyosis vulgaris. In some embodiments, ichthyosis vulgaris includes enhanced dermal texture of the palms, keratosis pilaris, and fine scaling most prominent on the lower abdomen, arms, and legs. In some embodiments, subjects may exhibit significant scaling. In some embodiments, subjects may exhibit enhanced dermal texture of the palms, keratosis pilaris, and possibly fine scaling. Accordingly, methods for treating subjects with ichthyosis vulgaris, or one or more symptoms or phenotypic characteristics thereof, are provided herein, comprising administering the subject an effective amount of one or more anti-KLK5 / KLK7 antibodies or one or more anti-KLK5 / KLK7+IL-4 bispecific antibodies.

[0231] Further embodiments of this disclosure relate to methods and compositions useful for treating psoriasis (e.g., anti-KLK5 / KLK7 antibodies or anti-KLK5 / KLK7 + IL-4 bispecific antibodies). Psoriasis (or psoriasis vulgaris) is a chronic inflammatory dermatitis. In some embodiments, psoriasis is characterized by red, scaly skin patches that may be found on the scalp, elbows, and / or knees of the subject. In some embodiments, psoriasis is associated with severe arthritis in the subject. In some embodiments, psoriasis-related lesions are caused by abnormal keratinocyte proliferation and infiltration of inflammatory cells into the dermis and epidermis. In some embodiments, the subject experiences the onset of psoriasis between the ages of 15 and 30. In some embodiments, psoriatic lesions are characterized by induration, scaling, and / or erythema of the skin, which may be accompanied by histological evidence of inflammation, abnormal keratinocyte proliferation / terminal differentiation, and / or dermal angiogenesis. In some embodiments, psoriatic inflammatory infiltrations may be prominent at the dermal-epidermal junction and contain activated T cells and antigen-presenting cells (APCs). In some embodiments, the presence of activated T cells and APCs in such infiltrations precedes the development of epidermal hyperplasia. In some embodiments, an increase in inflammatory cytokines is detectable in the lesions of psoriatic epidermis, which may lead to enhanced T cell activation and accelerated keratinocyte hyperplasia and differentiation. In some embodiments, methods for treating subjects having psoriasis are provided herein, comprising administering an effective amount of one or more anti-KLK5 / KLK7 antibodies or one or more anti-KLK5 / KLK7+IL-4 bispecific antibodies to the subject. In some embodiments, the subject is treated before the onset of epidermal hyperplasia. However, in some embodiments, the subject is treated after the onset of epidermal hyperplasia and accelerated keratinocyte differentiation.

[0232] Further aspects of this disclosure relate to methods and compositions useful for treating rosacea (e.g., anti-KLK5 / KLK7 antibodies or anti-KLK5 / KLK7+IL-4 bispecific antibodies). Rosacea is an inflammatory disease characterized by erythema, pustular papules, and / or telangiectasia. In some embodiments, subjects with rosacea express abnormally high levels of cathelicidine in their facial skin. In some embodiments, the proteolytic forms of cathelicidine peptides found in rosacea differ from those present in normal subjects. In some embodiments, methods for treating subjects with rosacea are provided herein, comprising administering an effective amount of one or more anti-KLK5 / KLK7 antibodies or one or more anti-KLK5 / KLK7+IL-4 bispecific antibodies to the subject.

[0233] Further aspects of this disclosure relate to methods for treating asthma-having subjects provided herein, comprising administering an effective amount of one or more antibodies disclosed herein (e.g., anti-KLK5 / KLK7 antibodies or anti-KLK5 / KLK7+IL-4 bispecific antibodies) to the subject. In some embodiments, methods for treating asthma-having subjects are provided herein, comprising administering an effective amount of one or more anti-KLK5 / KLK7 antibodies or one or more anti-KLK5 / KLK7+IL-4 bispecific antibodies. In some embodiments, the subject is asthma-having (e.g., a subject to be treated with a KLK5-targeted antibody provided herein). In some embodiments, asthma is selected from allergic asthma, aspirin-sensitive / exacerbating asthma, smoking-induced asthma, asthma uncontrolled with corticosteroids or other chronic asthma control agents, atopic asthma, bronchial obstruction-related asthma, etiology-related asthma, chronic asthma, corticosteroid-naive asthma, corticosteroid-refractory asthma, corticosteroid-resistant asthma, high eosinophil asthma, eosinophilic asthma, low eosinophil asthma, exercise-induced asthma, mild asthma, moderate to severe asthma, Netherton syndrome asthma, newly diagnosed and / or untreated asthma, non-allergic asthma, non-Th2-driven asthma, high periostin asthma, low periostin asthma, low Th2 asthma, type 2 (T2)-driven asthma, and low type 2 inflammatory asthma. In some embodiments, the subjects have atopic asthma or allergic asthma. In some embodiments, the subjects have aspirin-sensitive asthma or aspirin-exacerbated asthma. In some embodiments, the subjects have asthma associated with nonsteroidal anti-inflammatory drugs (NSAIDs). Thus, in some embodiments, the subjects have asthma induced by aspirin or a similar NSAID (e.g., recently ingested aspirin or a similar NSAID). In some embodiments, the subjects have bronchospasm that may or may not be characterized as asthma. For example, in some embodiments, the subjects have exercise-induced bronchospasm.

[0234] In some embodiments, the subjects have eosinophilic asthma. In some embodiments, the subjects have eosinophil-positive (EIP) asthma. In some embodiments, the subjects have eosinophil-negative (EIN) asthma. In some embodiments, the subjects have high eosinophil asthma (e.g., at least about 150, 200, 250, 300, 350, or 400 eosinophils / ml blood). In some embodiments, the subjects have low eosinophil asthma (e.g., less than about 150 eosinophils / μl or less than about 100 eosinophils / μl in the blood). These and further examples of asthma-related conditions treatable with the compositions provided herein (e.g., anti-KLK5 / KLK7 antibodies) are disclosed in WO2015 / 061441, METHODS OF DIAGNOSING AND TREATING EOSINOPHILIC DISORDERS, published April 30, 2015, and the relevant content of that publication is incorporated herein by reference.

[0235] In some embodiments, subjects have exercise-induced asthma, intermittent or exercise-induced, mild asthma, mild or corticosteroid-naive asthma, moderate to severe asthma, Netherton syndrome asthma, newly diagnosed untreated asthma, or severe asthma. In some embodiments, subjects have asthma that has never previously required or used the use of inhaled topical or systemic steroids (e.g., chronic use) to manage or control symptoms (e.g., cough, wheezing, shortness of breath / dyspnea, and chest pain).

[0236] In some embodiments, subjects have hypoperiostin asthma (e.g., having a serum periostin level of less than about 20 ng / mL). In some embodiments, subjects have hyperperiostin asthma (e.g., having a serum periostin level of at least about 20 ng / mL, 25 ng / mL, or 50 ng / mL). In some embodiments, subjects have non-allergic asthma (e.g., induced by, for example, infection with respiratory viruses (e.g., influenza, coronavirus, parainfluenza, rhinovirus, human metapneumovirus, and respiratory fusion viruses), or inhalation of irritants (air pollutants, smog, combustion particles (e.g., diesel particles), volatile chemicals, indoor and outdoor gases), or relatively cool, dry air, or not induced by such irritants). In some embodiments, subjects have asthma as a result of acute or chronic primary or indirect exposure to smoke (tobacco, cigars, pipes, or other combustion products), or inhalation or use of e-cigarettes (nicotine, cannabis, or other similar substances). In some embodiments, the subjects have persistent chronic severe asthma accompanied by acute events of potentially life-threatening symptom exacerbations (worsening or relapses).

[0237] In some embodiments, the subject has a type 2 (Th2) T helper lymphocyte or type 2 (Th2) high-level asthma condition. In some embodiments, the subject has Th2-induced asthma. For example, in some embodiments, Th2 cells and / or their secreted effector molecules mediate an immune response to an allergen, which is triggered by exposure to a specific allergen that results in allergic asthma in the subject. In some embodiments, the subject has activated Th2 cell-mediated asthma, which may be partially triggered by the secretion of interleukins, e.g., IL-4, IL-5, and IL-13. Therefore, in some embodiments, the KLK5 antibody (or other antibody) may be combined separately or in a multispecific antibody format with one or more antibodies targeting allergy-related targets such as IL-4 and IgE. Examples of such antibodies for the treatment of asthma include, but are not limited to, omalizumab (XOLAIR®) (targeting soluble IgE) and kilizumab (targeting membrane-bound IgE).

[0238] In other embodiments, the antibodies described herein are used to treat inflammatory disorders, infectious diseases, allergic diseases, and autoimmune disorders, but are not limited to those described below. In some embodiments, inflammatory disorders are selected from, but are not limited to, rosacea, prurigo nodosa, Crohn's disease, ankylosing spondylitis, ulcerative colitis, hidradenitis suppurativa, and uveitis, or one or more symptoms related to their barrier function. In some embodiments, allergic diseases are selected from, but are not limited to, eczema, atopic dermatitis, asthma, sinusitis, and eosinophilic esophagitis. In some embodiments, autoimmune diseases are selected from, but are not limited to, rheumatoid arthritis, psoriatic arthritis, juvenile idiopathic arthritis, Behçet's disease, and psoriasis vulgaris, or one or more symptoms related to their barrier function.

[0239] Determining whether the amount of antibody (e.g., anti-KLK5 / KLK7 antibody or anti-KLK5 / KLK7 + IL-4 bispecific antibody) has achieved a therapeutic effect will be obvious to those skilled in the art based on the teachings provided herein. The effective amount will vary depending on the specific condition being treated, the severity of the condition, the parameters of the individual patient, such as age, physical condition, size, sex, and weight, the duration of treatment, the nature of concurrent treatment (if any), the specific route of administration, and similar factors within the scope of the knowledge and expertise of the healthcare professional, as recognized by those skilled in the art. The specific administration plan used in the methods described herein, i.e., dose, timing, and repetitions, will depend on the specific subject and their medical history, as discussed herein.

[0240] Generally, empirical considerations, such as the time to maximum effect, half-life, and / or time above a certain concentration, will contribute to the determination of the dosage. For example, antibodies that are compatible with the human immune system, such as humanized or fully human antibodies, may be used to extend the antibody half-life and prevent the antibody from being attacked by the host immune system. Other reasons for adjusting the dose include differences in sex, age, individual response, polymorphism of the antibody target, and / or pharmacokinetic or pharmacodynamic responses driven by receptors involved in antibody clearance. The frequency of administration may be determined and adjusted throughout the course of treatment and is generally, but not necessarily, based on the treatment and / or suppression and / or improvement and / or delay of the target disease / impairment. Alternatively, sustained-release formulations of antibodies may be appropriate. Various formulations and devices for achieving sustained release are known in the art.

[0241] The frequency of administration may vary according to the claimed method. In some embodiments, the composition may be administered once. In some embodiments, the composition may be administered multiple times. In some embodiments, the frequency of administration may be weekly, every two weeks, every three weeks, every four weeks, every five weeks, every six weeks, every seven weeks, every eight weeks, every nine weeks, or every ten weeks; or once a month, every two months, or every three months, or at longer intervals. In some embodiments, the composition may be administered daily, every other week, weekly, every other month, monthly, or at any time interval that provides suitable (e.g., maximum) efficacy while minimizing safety risks to the subject. In general, efficacy and risks of treatment and safety may be monitored throughout the course of treatment.

[0242] In some embodiments, the subject may be administered one or more doses of the compositions provided herein (e.g., anti-KLK5 / KLK7 antibody or anti-KLK5 / KLK7 + IL-4 bispecific antibody) at intervals over a set period. In some cases, the periods during which the subject is administered the compositions at one or more intervals may be separated by periods during which the subject is not administered the compositions. In some embodiments, the relative duration of each period may depend on the subject's response to the treatment or the severity of the disease, or both, and / or may be determined based on the judgment of the physician administering the treatment.

[0243] In some embodiments, antibodies can be administered parenterally. For example, the parenterally administered composition may be administered topically, transmucosally, subcutaneously, intradermally, intravenously, intraperitoneally, intratumorally, intramuscularly, intraarticularly, intraarterially, or by infusion techniques.

[0244] In some embodiments, the antibody (e.g., anti-KLK5 / KLK7 antibody or anti-KLK5 / KLK7+IL-4 bispecific antibody) is administered intravenously. In some embodiments, the antibody (e.g., anti-KLK5 / KLK7 antibody or anti-KLK5 / KLK7+IL-4 bispecific antibody) is administered subcutaneously or topically.

[0245] For intravenous injection, water-soluble antibodies can be administered by drip infusion, thereby injecting a pharmaceutical preparation containing the antibody and physiologically acceptable excipients. Physiologically acceptable excipients may include, for example, 5% dextrose, 0.9% saline, Ringer's solution, or other suitable excipients. Other injectable compositions may contain various carriers, such as vegetable oil, dimethylactamide, dimethylformamide, ethyl lactate, ethyl carbonate, isopropyl myristate, ethanol, and polyols (glycerol, propylene glycol, liquid polyethylene glycol, etc.). In some cases, preparations, such as sterile preparations in a suitable soluble salt form of the antibody, can be dissolved and administered in pharmaceutical excipients such as water for injection, 0.9% saline, or 5% glucose solution.

[0246] In one embodiment, the antibody is administered via site-specific or targeted local delivery technology. Examples of site-specific or targeted local delivery technologies include various implantable, transdermal, or transmucosal depot sources of the antibody or local delivery system.

[0247] The anti-KLK5 / KLK7 antibodies or one or more anti-KLK5 / KLK7+IL-4 bispecific antibodies described herein and the treatment methods requiring them may be used in combination with other types of therapies for targeted diseases or disorders disclosed herein. In this regard, the antibody compositions and therapeutic agents may be administered simultaneously or sequentially. Such therapeutic agents may be administered simultaneously with or sequentially (in any order) the treatments according to this disclosure.

[0248] Accordingly, aspects of this disclosure relate to methods and compositions (e.g., anti-KLK5 / KLK7 antibodies or anti-KLK5 / KLK7+IL-4 bispecific antibodies). In some embodiments, the antibodies described herein may be administered as combination therapy (simultaneously or sequentially, e.g., over time). In some embodiments, the combination therapy involves administering one or more of the antibodies described herein (e.g., anti-KLK5 / KLK7 antibodies or anti-KLK5 / KLK7+IL-4 bispecific antibodies) and at least one additional therapeutic agent (e.g., therapeutic agents 1, 2, 3, 4, 5, 6, or 7). In some embodiments, one or more of the antibodies described herein and at least one additional therapeutic agent (e.g., therapeutic agents 1, 2, 3, 4, 5, 6, or 7) are administered together. In some embodiments, one or more of the antibodies described herein and at least one additional therapeutic agent (e.g., therapeutic agents 1, 2, 3, 4, 5, 6, or 7) are administered separately.

[0249] In some embodiments, additional therapeutic agents are anti-inflammatory agents. In some embodiments, the anti-inflammatory agents are selected from, but are not limited to, low-dose antibiotics, steroids, corticosteroids, tacrolimus, anti-IL4R antibodies (e.g., dupilumab), anti-IL-13 antibodies, TNF inhibitors (e.g., anti-TNF), IL-12 / 23 inhibitors, IL-17 inhibitors, and IL-4 receptor inhibitors, doxycycline, methotrexate, prednisone, cyclosporine, mycophenolate mofetil, dupilumab, certolizumab pegol, etanercept, adalimumab, infliximab, golimumab, ustekinumab, secukinumab, ixekizumab, brodalumab, abatacept, tidrakizumab-asmn, risankisumab-rzaa, and guselkumab. In some embodiments, the anti-inflammatory agent is administered orally. In some embodiments, the anti-inflammatory agent is administered topically. In some embodiments, the anti-inflammatory agent is administered by injection (e.g., intravenously, subcutaneously, or intramuscularly).

[0250] In some embodiments, the therapeutic combination comprises one or more of the antibodies described herein (e.g., anti-KLK5 / KLK7 antibodies or anti-KLK5 / KLK7+IL-4 bispecific antibodies) delivered together with one or more additional antibodies or fragments thereof (e.g., antibodies 1, 2, 3, 4, 5, 6, or 7). In some embodiments, the therapeutic combination comprising one or more of the antibodies described herein and one or more additional antibodies or fragments thereof (e.g., antibodies 1, 2, 3, 4, 5, 6, or 7) is delivered separately. In some embodiments, the therapeutic combination comprising one or more of the antibodies described herein and one or more additional antibodies or fragments thereof (e.g., antibodies 1, 2, 3, 4, 5, 6, or 7) is delivered together.

[0251] In some embodiments, a therapeutic combination comprising one or more antibodies described herein and one or more additional antibodies or fragments thereof (e.g., 1, 2, 3, 4, 5, 6, or 7 antibodies) is a multispecific antibody combination. In some embodiments, a multispecific antibody combination comprises a KLK5 / KLK7 antigen-specific binding site and one or more additional distinct antigen-specific binding sites from one or more additional antibodies. In some embodiments, a multispecific antibody comprises direct fusion or linkage of different antigen-specific binding sites. In some embodiments, additional antibodies or fragments thereof may be selected from, but are not limited to, anti-IL4R antibodies, anti-IL-13 antibodies, anti-TNF antibodies, anti-IL-12 / 23 antibodies, anti-IL-17 antibodies, doxycycline, dupilumab, certolizumab pegol, etanercept, adalimumab, infliximab, golimumab, ustekinumab, secukinumab, ixekizumab, brodalumab, abatacept, tildrakizumab-asmn, risankizumab-rzaa, and / or guselkumab.

[0252] In some embodiments, in non-limiting examples, in the treatment of rosacea, additional therapeutic agents may be selected from topical steroids; oral methotrexate; oral cyclosporine; and / or TNF inhibitors. In some embodiments, in non-limiting examples, in the treatment of atopic dermatitis, additional therapeutic agents may be selected from subcutaneous dupilumab and / or topical steroids.

[0253] Any anti-KLK5 / KLK7 antibody disclosed herein can be used to detect the presence of KLK5 and / or KLK7 in vitro or in vivo. Results obtained from such detection methods can be used for diagnostic purposes (e.g., diagnosis of diseases associated with KLK5 and / or KLK7) or for scientific research purposes (e.g., identification of novel KLK5-secreting cell types, study of the biological activity and / or regulation of secreted KLK5 and / or KLK7). For assay applications such as diagnostic applications, the anti-KLK5 / KLK7 antibodies or anti-KLK5 / KLK7+Th2 (e.g., anti-KLK5 / KLK7+IL-4) bispecific antibodies described herein may be conjugated with a detectable label (e.g., an imaging agent such as a contrast agent) for detecting the presence of KLK5 and / or KLK7 in vitro or in vivo.

[0254] As used herein, “conjugated” or “attached” means that two entities are associated, preferably with sufficient affinity to realize the therapeutic / diagnostic benefit of the association between the two entities. The association between two entities may occur directly or via a linker, such as a polymer linker. Conjugated or attached may include covalent or non-covalent bonds, as well as other forms of association, such as entrapments, e.g., micelles, where one entity is on or inside another, or where one or both entities are on or inside a third entity.

[0255] In other embodiments, the anti-KLK5 / KLK7 antibodies described herein can be conjugated to detectable labels, which are compounds capable of directly or indirectly emitting a detectable signal, thereby enabling detection, measurement, and / or identification of aptamers in vitro or in vivo. Examples of such “detectable labels” are intended to include, but are not limited to, fluorescent labels, chemiluminescent labels, colorimetric labels, enzyme markers, radioisotopes, and affinity tags such as biotin. Such labels can be conjugated to aptamers directly or indirectly by conventional methods.

[0256] The reporter agent may be a dye, such as a fluorophore, that is useful for detecting diseases mediated by KLK5 and / or KLK7-expressing cells in tissue samples.

[0257] For in vitro diagnostic assays, anti-KLK5 / KLK7 antibodies can be brought into contact with a sample suspected of containing KLK5 and / or KLK7, such as KLK5-expressing cells or a disease microenvironment containing soluble KLK5. The antibody and sample can be incubated for a suitable period under suitable conditions that allow the antibody to bind to the KLK5 antigen. Such interactions can then be detected by conventional methods, such as ELISA, histological staining, or FACS. For in vivo diagnostic assays, a suitable amount of anti-KLK5 / KLK7 antibody can be conjugated with a label (e.g., an imaging agent or contrast agent) and administered to the subject requiring testing. The presence of the labeled antibody can be detected by conventional methods based on the signal released from the label.

[0258] Anti-KLK5 / KLK7 antibodies can be used to perform scientific research assays to study the biological activity of KLK5 and / or KLK7, detect the presence of intracellular or extracellular KLK5 and / or KLK7, and / or control the effects of KLK5. For example, a suitable amount of anti-KLK5 / KLK7 can be contacted with a sample suspected of producing KLK5 and / or KLK7 (e.g., a new cell type not previously identified as a KLK5 and / or KLK7-producing cell). These cells are permeabilized before contact with the anti-KLK5 / KLK7 antibody. The antibody and sample can be incubated for a suitable period of time under suitable conditions that allow the antibody to bind to the KLK5 antigen. Such interactions can then be detected by conventional methods, such as ELISA, histological staining, or FACS.

[0259] VI. Kits for therapeutic and diagnostic use This disclosure also provides kits for therapeutic and diagnostic applications disclosed herein. Such kits may include one or more containers containing antibodies, for example, any of those described herein.

[0260] In some embodiments, the kit may include instructions for use in accordance with any of the methods described herein. The included instructions may include instructions for administering antibodies to treat, delay the onset of, or mitigate target diseases as described herein. The kit may further include instructions for selecting individuals suitable for treatment based on whether or not the individual has a target disease. In other embodiments, the instructions may include instructions for administering antibodies to individuals at risk of a target disease.

[0261] Instructions regarding the use of the described antibody generally include information on dosage, dosing schedule, and route of administration for the intended treatment. The container may be a unit dose, a bulk package (e.g., a multi-dose package), or a sub-unit dose. The instructions provided in the kits of the present invention are typically written instructions on a label or an accompanying document (e.g., a sheet of paper included in the kit), but machine-readable instructions (e.g., instructions stored on a magnetic or optical storage disk) are also acceptable.

[0262] The label or the accompanying document indicates that the composition is used to treat, delay the onset of, and / or alleviate a disease or disorder. Instructions for performing any of the methods described herein may be provided.

[0263] The kits of the present invention are contained in suitable packaging. Suitable packaging includes, but is not limited to, vials, bottles, jars, flexible packaging (e.g., shielded mylar or plastic bags), etc.

[0264] Packaging for use in combination with specific devices, such as infusion devices, such as mini pumps, is also contemplated. The kit may have a sterile access port (e.g., the container may be an intravenous solution bag or vial having a stopper that can be punctured by a hypodermic needle). Also, the container may have a sterile access port (e.g., the container may be an intravenous solution bag or vial having a stopper that can be punctured by a hypodermic needle). At least one active agent in the composition is the antibody described herein.

[0265] The kit may optionally provide additional components such as buffers and interpretive information. Generally, the kit includes a container and a label or accompanying document(s) on or associated with the container. In some embodiments, the present invention provides a pharmaceutical product comprising the contents of the above-described kit.

[0266] Kits for use in detecting target proteins (e.g., KLK5 and / or KLK7, or KLK5 and KLK7 + IL-4) in a sample are also provided herein. Such kits may include any of the antibodies described herein. In some examples, the antibodies may be conjugated with detectable labels described herein. As used herein, “conjugated” or “attached” means that two entities are associated, preferably with sufficient affinity to realize the therapeutic / diagnostic benefit of the association between the two entities. The association between the two entities may occur directly or via a linker such as a polymer linker. Conjugated or attached can include covalent or non-covalent bonds, as well as other forms of association, such as entrapment, e.g., micelles, where one entity is on or inside another, or where one or both entities are on or inside a third entity.

[0267] Alternatively or additionally, the kit may include a second antibody that can bind to the antibody described herein. The kit may further include instructions for using the antibody to detect a target protein (e.g., KLK5 and / or KLK7, or KLK5 and KLK7 + IL-4). [Examples]

[0268] Example 1: Generation and selection of dual inhibitory anti-KLK5 / KLK7 antibodies (i) Affinity Antibody binding kinetics experiments were performed via Biacore to screen for anti-KLK5 / KLK7 antibodies with high affinity for each target. All screening assays were performed at 25°C. The running buffer used was 20 mM HEPES, 300 mM NaCl, 0.01% Tween®-20, pH 7.5. Antibody (1 ug / mL) was captured on Fc2-4 of a Series S Protein A sensor chip (Cytiva) at a flow rate of 10 ul / mL for 30 seconds. Reaction kinetics were performed in single-cycle reaction mode using a series of 4-5 concentrations at maximum concentrations of 25-100 nM and 4-fold serial dilutions. The addition time was typically 300 seconds, and the dissociation time ranged from 1800-3600 seconds. A typical flow rate of 30 ul / mL was used. The sensor chip was regenerated using 10 mM glycine, pH 1.5, with an addition time of 30 seconds and a flow rate of 50 ul / mL. The results indicate that the dual inhibitory antibodies listed in Table 1a exhibited binding specificity for both KLK5 and KLK7. A summary of the binding kinetics of the anti-KLK5 antibody is provided in Table 4 below. [Table 4]

[0269] (ii) Protease inhibition assay To evaluate the ability of the antibodies listed in Table 1a to carefully inhibit KLK5 and / or KLK7 protease activity, 1.5 nM human or mouse KLK5 or 0.5 nM human or mouse KLK7 were prepared in assay buffer (0.1 M NaH2PO4, pH=7.5 for KLK5, and 50 mM Tris, 150 mM NaCl, pH=7.5 for KLK7). The antibodies, diluted in PBS to the test concentration, were added. Then the substrates were added (50 μM BOC-Val-Pro-Arg-AMC for KLK5, and 30 μM KHLF-AMC for KLK7). The fluorescence signal was then measured minute by minute at room temperature for 30 minutes.

[0270] The results, as shown in Table 4, demonstrate that anti-KLK5 / KLK7 antibodies can inhibit KLK5 and KLK7 protease activity.

[0271] Example 2: Production of anti-KLK5 / KLK7+IL-4 targeted bispecific antibodies Experiments were conducted to demonstrate that inhibiting KLK5 / KLK7 using a dual inhibitory anti-KLK5 / KLK7 antibody in combination with an anti-IL-4R antibody had a more beneficial effect than either antibody alone in an Nc / Nga mouse model. Topical application of house dust mite (HDM) allergen induced mouse atopic dermatitis in Nc / Nga mice (a strain with an endogenous barrier defect due to reduced ceramide expression). Animals aged 8-10 weeks received Biostir-AD ointment (120 mg / mouse) containing a mite allergen derived from Dermatophagoides farina, applied twice a week for two weeks to both ears and dorsal skin areas (including the neck), for a total of 480 mg per mouse. 150 μL of SDS was applied 2 hours before Biostir-AD application from the second treatment (day 4). The vehicle or test product (control IgG, KLK5-KLK7-dual Ab4, KLK5 / 7 inhibitor, anti-IL-4R inhibitory antibody, or a combination of KLK5 / KLK7-dual-Ab4 and anti-IL-4R inhibitory antibody) was administered intraperitoneally (IP) at a dose of 30 mg / kg three times / week for a total of nine doses, starting on day -6. The anti-IL-4R inhibitory antibody inhibits IL-4 signaling. The reference compound, tacrolimus ointment, was applied topically (TOP) once daily (QD) for a total of 21 days (QD x 21), starting on day -6 (QD x 21), and applied one hour after Biostir-AD application on the Biostir-AD challenge day. The model was evaluated for macroscopic skin lesion assessment and ear thickness measurement on day 15. Ear thickness was measured using a Dyer model micrometer gauge. The severity of skin lesions (four parameters: erythema / hemorrhage, edema, peeling / erosion, and scaling / dryness) is assessed by signs on the ears, neck, and dorsal skin. The overall clinical skin severity score is defined as the sum of the individual scores (0: none, 1: mild, 2: moderate, 3: severe).

[0272] The results showed that combined inhibition of KLK5 / KLK7 and IL-4 signaling had an enhanced effect in reducing ear thickness and clinical skin score compared to inhibiting each antibody alone.

[0273] Therefore, it is advantageous to generate a bispecific antibody that inhibits KLK5 / KLK7 activity and IL-4 signaling with a single molecule. For example, a bispecific antibody having one arm with an antigen-specific binding site of the anti-KLK5 / KLK7 antibody described in Table 1a and the other arm with an antigen-specific binding site of the IL-4 targeted antibody described in Table 2 is suitable for inhibiting KLK5 / KLK7 and IL-4 signaling via a single molecule.

[0274] Other Embodiments All features disclosed herein may be combined in any combination. Each feature disclosed herein may be replaced by an alternative feature that serves the same, equivalent, or similar purpose. Thus, unless otherwise expressly indicated, each disclosed feature is merely an example of a general set of equivalent or similar features.

[0275] From the above description, those skilled in the art will be able to easily grasp the essential features of the present invention and adapt it to various uses and situations by making various changes and modifications without departing from the spirit and scope of this disclosure.

[0276] Equivalents and Scope In the claims, articles such as “a,” “an,” and “the” may mean one or more unless otherwise indicated or the context makes it clear. A claim or description containing “or” between one or more members of a group is deemed satisfied unless otherwise indicated or the context makes it clear that one, more or all of the members of that group are present, adopted or otherwise related in a given product or process. The present invention includes embodiments in which exactly one member of a group is present, adopted or otherwise related in a given product or process. The present invention includes embodiments in which more or all of the members of a group are present, adopted or otherwise related in a given product or process.

[0277] Furthermore, the present invention encompasses all variations, combinations, and rearrangements in which one or more limitations, elements, clauses, and descriptive terms from one or more of the enumerated claims are introduced into another claim. For example, any claim dependent on another claim can be modified to include one or more limitations found in any other claim dependent on the same basic claim. If elements are presented as a list, for example in Markush group format, each subgroup of the elements is also disclosed, and any element(s) can be removed from that group. In general, where the present invention or an aspect of the present invention is referred to as including certain elements and / or features, it should be understood that a particular embodiment or aspect of the present invention consists of, or is essentially derived from, such elements and / or features. For brevity, these embodiments are not specifically described herein by these words. As used herein and in the claims, the phrase “and / or” should be understood to mean “either or both” of such associated elements, i.e., elements that exist together in some cases and disjunct in others. Multiple elements listed with “and / or” should be interpreted in the same manner, i.e., “one or more” of the elements thus associated. Other elements other than those specifically identified by the “and / or” clause may exist at their discretion, whether or not they are related to the specifically identified elements. Thus, as a non-restrictive example, when used in conjunction with non-restrictive language such as “comprising,” a reference to “A and / or B” may refer to A only in one embodiment (optionally including elements other than B); B only in another embodiment (optionally including elements other than A); and still in yet another embodiment to both A and B (optionally including other elements).

[0278] As used herein and in the claims, “or” should be understood to have the same meaning as “and / or” as defined above. For example, when separating items in a list, “or” or “and / or” should be interpreted as inclusive, that is, including at least one of several elements or lists of elements and additional items not listed by choice, but also including multiple items. Only terms that indicate the exact opposite, such as “one of” or “exactly one of” or, as used in the claims, “consisting of,” would refer to including exactly one element of several elements or lists of elements. In general, as used herein, the term “or” should be interpreted as indicating exclusive substitutes (i.e., “one or the other, but not both”) only when preceded by exclusive terms such as “either,” “one of,” “one of” or “exactly one of.” As used in the claims, “essentially consisting of” should have the usual meaning as used in the field of patent law.

[0279] As used in this specification and the claims, when referring to a list of one or more elements, the phrase "at least one" means at least one element selected from any one or more of the elements in the list of elements, and does not necessarily have to include at least one of each and every element specifically listed in the list of elements, and is not to exclude any combination of elements in the list of elements. It should be understood that this definition also allows for the optional presence of elements other than those specifically identified in the list of elements referred to by the phrase "at least one", whether or not related to the specifically identified elements. Thus, by way of non-limiting example, "at least one of A and B" (or equivalently, "at least one of A or B" or equivalently "at least one of A and / or B") can, in one embodiment, refer to at least one A, optionally including two or more, with B not present (optionally including elements other than B); in another embodiment, it can refer to at least one B, optionally including two or more, with A not present (optionally including elements other than A); in yet another embodiment, it can refer to at least one A, optionally including two or more, and at least one B, optionally including two or more (optionally including other elements); and so on.

[0280] Also, it should be understood that, unless expressly indicated to the contrary, in any method claimed herein that includes a plurality of steps or acts, the order of the method steps or acts is not necessarily limited to the order in which the method steps or acts are described.

[0281] As with the above specification, all transitional phrases in the claims, such as “comprising,” “including,” “possessing,” “having,” “containing,” “accompanying,” “holding,” and “composed of,” should be understood to be non-restrictive, meaning they include it but are not limited to it. Only the transitional phrases “essentially consisting of” and “essentially consisting of” are considered restrictive or semi-restrictive transitional phrases, respectively, as described in Section 2111.03 of the U.S. Patent and Trademark Office's Examination Manual. Embodiments described herein using non-restrictive transitional phrases (e.g., “including”) should be understood to also be intended in alternative embodiments to be features described by non-restrictive transitional phrases, such as “~consisting of” and “~essentially consisting of.” For example, if this application describes “a composition comprising A and B,” this application also intends alternative embodiments “a composition comprising A and B” and “a composition essentially consisting of A and B.”

[0282] If a range is given, it includes the endpoints. Furthermore, unless otherwise indicated or otherwise evident from the context and the understanding of those skilled in the art, the values ​​expressed as a range may be any specific value or subrange within the range specified in various embodiments of the invention, up to one-tenth of the lower limit of the range, unless explicitly indicated otherwise in the context.

[0283] This application references various published patents, published patent applications, academic papers, and other publications, all of which are incorporated herein by reference. In the event of any conflict between any of the incorporated references and this specification, this specification shall prevail. In addition, any particular embodiment of the Invention that falls within the scope of prior art may be clearly excluded from any one or more of the claims. Such embodiments are considered to be known to those skilled in the art and may be excluded even if such exclusion is not expressly stated herein. Any particular embodiment of the Invention may be excluded from any claim for any reason, whether or not it relates to the existence of prior art.

[0284] Those skilled in the art will be able to recognize or confirm many equivalents to the specific embodiments described herein by simply using conventional experiments. The scope of the embodiments of the present invention described herein is not intended to be limited to the embodiments for carrying out the invention described above, but is as set out in the appended claims. Those skilled in the art will understand that various changes and modifications may be made to this specification without departing from the spirit or scope of the invention as defined in the following claims.

[0285] The enumeration of lists of chemical groups in any definition of a variable herein includes the definition of the variable as any single group or as a combination of the enumerated groups. The enumeration of embodiments relating to a variable herein includes embodiments as any single embodiment or embodiments in combination with any other embodiments or parts thereof. The enumeration of embodiments herein includes embodiments as any single embodiment or embodiments in combination with any other embodiments or parts thereof.

Claims

1. A bispecific antibody comprising an antigen-specific binding site that specifically binds to KLK5 and KLK7, and an antigen-specific binding site that specifically binds to interleukin (IL)-4 or its receptor.

2. The bispecific antibody according to claim 1, wherein the antigen-specific binding sites that specifically bind to KLK5 and KLK7 bind to the active sites of KLK5 and KLK7 and inhibit the activity of the enzymes.

3. The bispecific antibody according to claim 1 or 2, wherein the antigen-specific binding site that specifically binds to KLK5 and KLK7 comprises one of the antibodies listed in Tables 1a and 1b: HC CDR1, HC CDR2, HC CDR3, LC CDR1, LC CDR2, and / or LC CDR3.

4. The antigen-specific binding sites that specifically bind to KLK5 and KLK7 are (a) HC CDR1, HC CDR2, and HC CDR3 heavy chain variable domains having the amino acid sequence of SEQ ID NO: 7, and LC CDR1, LC CDR2, and LC CDR3 light chain variable domains having the amino acid sequence of SEQ ID NO: 8, (b) HC CDR1, HC CDR2, and HC CDR3 heavy chain variable domains having the amino acid sequence of SEQ ID NO: 13, and LC CDR1, LC CDR2, and LC CDR3 light chain variable domains having the amino acid sequence of SEQ ID NO: 14, (c) HC CDR1, HC CDR2, and HC CDR3 heavy chain variable domains having the amino acid sequence of SEQ ID NO: 17, and LC CDR1, LC CDR2, and LC CDR3 light chain variable domains having the amino acid sequence of SEQ ID NO: 14, or (d) A bispecific antibody according to any one of claims 1 to 3, comprising HC CDR1, HC CDR2, and HC CDR3 heavy chain variable domains having the amino acid sequence of SEQ ID NO: 21, and LC CDR1, LC CDR2, and LC CDR3 light chain variable domains having the amino acid sequence of SEQ ID NO:

14.

5. The antigen-specific binding sites that specifically bind to KLK5 and KLK7 are (a) HC CDR1 having the amino acid sequence of SEQ ID NO: 1, HC CDR2 having the amino acid sequence of SEQ ID NO: 2, HC CDR3 having the amino acid sequence of SEQ ID NO: 3, LC CDR1 having the amino acid sequence of SEQ ID NO: 4, LC CDR2 having the amino acid sequence of SEQ ID NO: 5, and LC CDR3 having the amino acid sequence of SEQ ID NO: 6 (b) HC CDR1 having the amino acid sequence of SEQ ID NO: 9, HC CDR2 having the amino acid sequence of SEQ ID NO: 10, HC CDR3 having the amino acid sequence of SEQ ID NO: 11, LC CDR1 having the amino acid sequence of SEQ ID NO: 4, LC CDR2 having the amino acid sequence of SEQ ID NO: 5, and LC CDR3 having the amino acid sequence of SEQ ID NO: 12 (c) HC CDR1 having the amino acid sequence of SEQ ID NO: 9, HC CDR2 having the amino acid sequence of SEQ ID NO: 15, HC CDR3 having the amino acid sequence of SEQ ID NO: 16, LC CDR1 having the amino acid sequence of SEQ ID NO: 4, LC CDR2 having the amino acid sequence of SEQ ID NO: 5, and LC CDR3 having the amino acid sequence of SEQ ID NO: 12, or (d) A bispecific antibody according to any one of claims 1 to 4, comprising HC CDR1 having the amino acid sequence of SEQ ID NO: 18, HC CDR2 having the amino acid sequence of SEQ ID NO: 19, HC CDR3 having the amino acid sequence of SEQ ID NO: 20, LC CDR1 having the amino acid sequence of SEQ ID NO: 4, LC CDR2 having the amino acid sequence of SEQ ID NO: 5, and LC CDR3 having the amino acid sequence of SEQ ID NO:

12.

6. The bispecific antibody according to any one of claims 1 to 5, wherein the antigen-specific binding site that specifically binds to KLK5 and KLK7 is the VH and / or VL of any one of the antibodies listed in Table 1a.

7. The antigen-specific binding sites that specifically bind to KLK5 and KLK7 are (a) VH containing the amino acid sequence of SEQ ID NO: 7, and VL containing the amino acid sequence of SEQ ID NO: 8, (b) VH containing the amino acid sequence of SEQ ID NO: 13, and VL containing the amino acid sequence of SEQ ID NO: 14, (c) VH containing the amino acid sequence of SEQ ID NO: 17, and VL containing the amino acid sequence of SEQ ID NO: 14, (d) A bispecific antibody according to any one of claims 1 to 6, comprising VH containing the amino acid sequence of SEQ ID NO: 21 and VL containing the amino acid sequence of SEQ ID NO:

14.

8. A bispecific antibody according to any one of claims 1 to 7, wherein the antigen-specific binding site that specifically binds to IL-4 or its receptor comprises one of the antibodies listed in Table 2: HC CDR1, HC CDR2, HC CDR3, LC CDR1, LC CDR2, and / or LC CDR3.

9. The antigen-specific binding site that specifically binds to IL-4 or IL-4R is (a) HC CDR1, HC CDR2, and HC CDR3 heavy chain variable domains having the amino acid sequence of SEQ ID NO: 154, and LC CDR1, LC CDR2, and LC CDR3 light chain variable domains having the amino acid sequence of SEQ ID NO: 155, (b) HC CDR1, HC CDR2, and HC CDR3 heavy chain variable domains having the amino acid sequence of SEQ ID NO: 174, and LC CDR1, LC CDR2, and LC CDR3 light chain variable domains having the amino acid sequence of SEQ ID NO: 175, (c) HC CDR1, HC CDR2, and HC CDR3 heavy chain variable domains having the amino acid sequence of SEQ ID NO: 194, and LC CDR1, LC CDR2, and LC CDR3 light chain variable domains having the amino acid sequence of SEQ ID NO: 195, (d) HC CDR1, HC CDR2, and HC CDR3 heavy chain variable domains having the amino acid sequence of SEQ ID NO: 214, and LC CDR1, LC CDR2, and LC CDR3 light chain variable domains having the amino acid sequence of SEQ ID NO: 215, (e) HC CDR1, HC CDR2, and HC CDR3 heavy chain variable domains having the amino acid sequence of SEQ ID NO: 234, and LC CDR1, LC CDR2, and LC CDR3 light chain variable domains having the amino acid sequence of SEQ ID NO: 235, or (f) The bispecific antibody according to claim 8, comprising HC CDR1, HC CDR2, and HC CDR3 heavy chain variable domains having the amino acid sequence of SEQ ID NO: 254, and LC CDR1, LC CDR2, and LC CDR3 light chain variable domains having the amino acid sequence of SEQ ID NO:

255.

10. The antigen-specific binding site that specifically binds to IL-4 or IL-4R is (a) HC CDR1 having the amino acid sequence of SEQ ID NO: 156, HC CDR2 having the amino acid sequence of SEQ ID NO: 157, HC CDR3 having the amino acid sequence of SEQ ID NO: 158, LC CDR1 having the amino acid sequence of SEQ ID NO: 159, LC CDR2 having the amino acid sequence of SEQ ID NO: 160, and LC CDR3 having the amino acid sequence of SEQ ID NO: 161 (b) HC CDR1 having the amino acid sequence of SEQ ID NO: 162, HC CDR2 having the amino acid sequence of SEQ ID NO: 163, HC CDR3 having the amino acid sequence of SEQ ID NO: 164, LC CDR1 having the amino acid sequence of SEQ ID NO: 165, LC CDR2 having the amino acid sequence of LGS, and LC CDR3 having the amino acid sequence of SEQ ID NO: 167 (c) HC CDR1 having the amino acid sequence of SEQ ID NO: 168, HC CDR2 having the amino acid sequence of SEQ ID NO: 169, HC CDR3 having the amino acid sequence of SEQ ID NO: 170, LC CDR1 having the amino acid sequence of SEQ ID NO: 171, LC CDR2 having the amino acid sequence of SEQ ID NO: 172, and LC CDR3 having the amino acid sequence of SEQ ID NO: 173, or (d) HC CDR1 having the amino acid sequence of SEQ ID NO: 176, HC CDR2 having the amino acid sequence of SEQ ID NO: 177, HC CDR3 having the amino acid sequence of SEQ ID NO: 178, LC CDR1 having the amino acid sequence of SEQ ID NO: 179, LC CDR2 having the amino acid sequence of SEQ ID NO: 180, and LC CDR3 having the amino acid sequence of SEQ ID NO: 181 (e) HC CDR1 having the amino acid sequence of SEQ ID NO: 182, HC CDR2 having the amino acid sequence of SEQ ID NO: 183, HC CDR3 having the amino acid sequence of SEQ ID NO: 184, LC CDR1 having the amino acid sequence of SEQ ID NO: 185, LC CDR2 having the amino acid sequence of AAS, and LC CDR3 having the amino acid sequence of SEQ ID NO: 187 (f) HC CDR1 having the amino acid sequence of SEQ ID NO: 188, HC CDR2 having the amino acid sequence of SEQ ID NO: 189, HC CDR3 having the amino acid sequence of SEQ ID NO: 190, LC CDR1 having the amino acid sequence of SEQ ID NO: 191, LC CDR2 having the amino acid sequence of SEQ ID NO: 192, and LC CDR3 having the amino acid sequence of SEQ ID NO: 193 (g) HC CDR1 having the amino acid sequence of SEQ ID NO: 196, HC CDR2 having the amino acid sequence of SEQ ID NO: 197, HC CDR3 having the amino acid sequence of SEQ ID NO: 198, LC CDR1 having the amino acid sequence of SEQ ID NO: 199, LC CDR2 having the amino acid sequence of SEQ ID NO: 200, and LC CDR3 having the amino acid sequence of SEQ ID NO:

201. (h) HC CDR1 having the amino acid sequence of SEQ ID NO: 202, HC CDR2 having the amino acid sequence of SEQ ID NO: 203, HC CDR3 having the amino acid sequence of SEQ ID NO: 204, LC CDR1 having the amino acid sequence of SEQ ID NO: 205, LC CDR2 having the amino acid sequence of SEQ ID NO: 206, and LC CDR3 having the amino acid sequence of SEQ ID NO: 207, (i) HC CDR1 having the amino acid sequence of SEQ ID NO: 208, HC CDR2 having the amino acid sequence of SEQ ID NO: 209, HC CDR3 having the amino acid sequence of SEQ ID NO: 210, LC CDR1 having the amino acid sequence of SEQ ID NO: 211, LC CDR2 having the amino acid sequence of GAS, and LC CDR3 having the amino acid sequence of SEQ ID NO: 213 (j) HC CDR1 having the amino acid sequence of SEQ ID NO: 216, HC CDR2 having the amino acid sequence of SEQ ID NO: 217, HC CDR3 having the amino acid sequence of SEQ ID NO: 218, LC CDR1 having the amino acid sequence of SEQ ID NO: 219, LC CDR2 having the amino acid sequence of SEQ ID NO: 220, and LC CDR3 having the amino acid sequence of SEQ ID NO:

221. (k) HC CDR1 having the amino acid sequence of SEQ ID NO: 222, HC CDR2 having the amino acid sequence of SEQ ID NO: 223, HC CDR3 having the amino acid sequence of SEQ ID NO: 224, LC CDR1 having the amino acid sequence of SEQ ID NO: 225, LC CDR2 having the amino acid sequence of YTS, and LC CDR3 having the amino acid sequence of SEQ ID NO:

227. (l) HC CDR1 having the amino acid sequence of SEQ ID NO: 228, HC CDR2 having the amino acid sequence of SEQ ID NO: 229, HC CDR3 having the amino acid sequence of SEQ ID NO: 230, LC CDR1 having the amino acid sequence of SEQ ID NO: 231, LC CDR2 having the amino acid sequence of SEQ ID NO: 232, and LC CDR3 having the amino acid sequence of SEQ ID NO:

233. (m) HC CDR1 having the amino acid sequence of SEQ ID NO: 236, HC CDR2 having the amino acid sequence of SEQ ID NO: 237, HC CDR3 having the amino acid sequence of SEQ ID NO: 238, LC CDR1 having the amino acid sequence of SEQ ID NO: 239, LC CDR2 having the amino acid sequence of SEQ ID NO: 240, and LC CDR3 having the amino acid sequence of SEQ ID NO:

241. (n) HC CDR1 having the amino acid sequence of SEQ ID NO: 242, HC CDR2 having the amino acid sequence of SEQ ID NO: 243, HC CDR3 having the amino acid sequence of SEQ ID NO: 244, LC CDR1 having the amino acid sequence of SEQ ID NO: 245, LC CDR2 having the amino acid sequence of SEQ ID NO: 246, and LC CDR3 having the amino acid sequence of SEQ ID NO:

247. (o) HC CDR1 having the amino acid sequence of SEQ ID NO: 248, HC CDR2 having the amino acid sequence of SEQ ID NO: 249, HC CDR3 having the amino acid sequence of SEQ ID NO: 250, LC CDR1 having the amino acid sequence of SEQ ID NO: 251, LC CDR2 having the amino acid sequence of SEQ ID NO: 252, and LC CDR3 having the amino acid sequence of SEQ ID NO: 253 (p) HC CDR1 having the amino acid sequence of SEQ ID NO: 256, HC CDR2 having the amino acid sequence of SEQ ID NO: 257, HC CDR3 having the amino acid sequence of SEQ ID NO: 258, LC CDR1 having the amino acid sequence of SEQ ID NO: 259, LC CDR2 having the amino acid sequence of SEQ ID NO: 260, and LC CDR3 having the amino acid sequence of SEQ ID NO:

261. (q) HC CDR1 having the amino acid sequence of SEQ ID NO: 262, HC CDR2 having the amino acid sequence of SEQ ID NO: 263, HC CDR3 having the amino acid sequence of SEQ ID NO: 264, LC CDR1 having the amino acid sequence of SEQ ID NO: 265, LC CDR2 having the amino acid sequence of SEQ ID NO: 267, and LC CDR3 having the amino acid sequence of SEQ ID NO: 268, or (r) A bispecific antibody according to claim 8 or 9, comprising HC CDR1 having the amino acid sequence of SEQ ID NO: 269, HC CDR2 having the amino acid sequence of SEQ ID NO: 270, HC CDR3 having the amino acid sequence of SEQ ID NO: 271, LC CDR1 having the amino acid sequence of SEQ ID NO: 272, LC CDR2 having the amino acid sequence of SAS, and LC CDR3 having the amino acid sequence of SEQ ID NO:

274.

11. The antigen-specific binding site that specifically binds to IL-4 or IL-4R is (a) VH containing the amino acid sequence of SEQ ID NO: 154, and VL containing the amino acid sequence of SEQ ID NO: 155, (b) VH containing the amino acid sequence of SEQ ID NO: 174, and VL containing the amino acid sequence of SEQ ID NO: 175, (c) VH containing the amino acid sequence of SEQ ID NO: 194, and VL containing the amino acid sequence of SEQ ID NO: 195, (d) VH containing the amino acid sequence of SEQ ID NO: 214, and VL containing the amino acid sequence of SEQ ID NO: 215, (e) VH containing the amino acid sequence of SEQ ID NO: 234, and VL containing the amino acid sequence of SEQ ID NO: 235, (f) A bispecific antibody according to any one of claims 8 to 10, comprising VH containing the amino acid sequence of SEQ ID NO: 254 and VL containing the amino acid sequence of SEQ ID NO:

255.

12. The bispecific antibody according to any one of claims 1 to 5, wherein the antigen-specific binding site that specifically binds to IL-4 or its receptor is the VH and / or VL of any one of the antibodies in Table 2.

13. The bispecific antibody according to any one of claims 1 to 12, wherein the bispecific antibody maintains a binding affinity for KLK5 and KLK7 that is 20% or less lower than that of an anti-KLK5 / KLK7 antibody containing the same CDR and / or VH / VL as the bispecific antibody.

14. The bispecific antibody according to any one of claims 1 to 12, wherein the bispecific antibody maintains a binding affinity for IL-4 that is 20% or less lower than that of an anti-IL-4 antibody containing the same CDR and / or VH / VL as the bispecific antibody.

15. The bispecific antibody according to any one of claims 1 to 14, wherein the bispecific antibody retains at least 80% of the inhibitory activity against KLK5 and KLK7 compared to an anti-KLK5 / KLK7 antibody containing the same CDR and / or VH / VL as the bispecific antibody.

16. The bispecific antibody according to any one of claims 1 to 15, wherein the bispecific antibody retains at least 80% of the inhibitory activity on IL-4 signaling compared to an anti-IL-4 antibody containing the same CDR and / or VH / VL as the bispecific antibody.

17. A composition comprising a bispecific antibody according to any one of claims 1 to 16 and an acceptable carrier.

18. A method comprising administering a bispecific antibody according to any one of claims 1 to 16 or the composition according to claim 17 to a target.

19. The method according to claim 17, wherein the subject has a skin barrier defect.

20. A method for treating a skin barrier defect, comprising administering an effective amount to a target of a bispecific antibody according to any one of claims 1 to 16 or a composition according to claim 17.

21. The method according to claim 19 or 20, wherein the skin barrier defect is associated with Netherton syndrome, atopic dermatitis, eosinophilic esophagitis, prurigo nodosa, chronic pruritus of unknown cause (CPUO), dry skin, asthma (especially KLK5), ichthyosis vulgaris, or itching or chronic itching.

22. The method according to any one of claims 18 to 21, wherein the subject has atopic dermatitis.

23. The method according to any one of claims 18 to 22, wherein the administration reduces ear thickness by more than 30% compared to a subject administered with an anti-IL-4 antibody containing the same CDR and / or VH / VL as the bispecific antibody.

24. The method according to any one of claims 18 to 23, wherein the administration reduces ear thickness by more than 30% compared to a subject administered with an anti-IL-4 antibody containing the same CDR and / or VH / VL as the bispecific antibody.

25. The method according to any one of claims 18 to 24, wherein the administration reduces skin erythema / hemorrhage by more than 30% compared to subjects administered with an anti-IL-4 antibody containing the same CDR and / or VH / VL as the bispecific antibody.

26. The method according to any one of claims 18 to 25, wherein the administration reduces skin erythema / hemorrhage by more than 30% compared to subjects administered with an anti-IL-4 antibody containing the same CDR and / or VH / VL as the bispecific antibody.

27. The method according to any one of claims 18 to 26, wherein the administration reduces skin erythema / hemorrhage by more than 30% compared to subjects administered with an anti-IL-4 antibody containing the same CDR and / or VH / VL as the bispecific antibody.

28. The method according to any one of claims 18 to 27, wherein the administration reduces skin peeling / erosion by more than 30% compared to subjects administered with an anti-IL-4 antibody containing the same CDR and / or VH / VL as the bispecific antibody.

29. The method according to any one of claims 18 to 28, wherein the administration reduces skin scaling / dryness by more than 30% compared to subjects administered with an anti-IL-4 antibody containing the same CDR and / or VH / VL as the bispecific antibody.

30. The method according to any one of claims 18 to 29, wherein the administration reduces skin scaling / dryness by more than 30% compared to subjects administered with an anti-IL-4 antibody containing the same CDR and / or VH / VL as the bispecific antibody.

31. The method according to any one of claims 18 to 30, wherein the administration reduces skin edema by more than 30% compared to subjects administered with an anti-IL-4 antibody containing the same CDR and / or VH / VL as the bispecific antibody.

32. The method according to any one of claims 18 to 31, wherein the administration reduces skin edema by more than 30% compared to subjects administered with an anti-IL-4 antibody containing the same CDR and / or VH / VL as the bispecific antibody.

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