Antibodies against canine interleukin-4 receptor alpha

Abstract

To provide an isolated mammalian antibody or an antigen binding fragment thereof that specifically binds to canine interleukin-4 receptor α (IL-4Rα).SOLUTION: The present invention discloses antibodies and blocking antibodies to canine IL-4 receptor alpha that have specific sequences and a high binding affinity for canine IL-4 receptor α. The present invention also discloses the use of the antibodies of the present invention in the treatment of atopic dermatitis in dogs. The present invention further discloses unique epitopes that bind to the antibodies to canine IL-4 receptor alpha.SELECTED DRAWING: Figure 1

Description

[Technical Field]

[0001] Cross-references to related applications This application is U.S. Provisional Patent Application No. 62 / 142,108, filed on April 2, 2015. Application No. 62 / 269,486 dated December 18, 2015 and March 18, 2016 Attachment application No. 62 / 310,250 (all of the contents of which are incorporated herein by reference) (to be included) Although claiming priority under 35 USC §119(e) That is the case.

[0002] Field of Invention This invention provides high binding affinity and specific distribution to the canine IL-4 receptor alpha. Antibodies against canine IL-4 receptor alpha (to canine IL-4 receptor alpha) The present invention relates to several that can block the binding of canine IL-4. This invention relates to a specific epitope that binds to antibodies against receptor alpha. The present invention also relates to This invention relates to the use of antibodies and epitopes in the treatment of atopic dermatitis. [Background technology]

[0003] The immune system works collaboratively to defend the host from infections and cancer, with commensal and recurrent immune systems. It includes a network of circulating specialized cells. The ability of the immune system to perform this function is due to white blood cells. It is largely dependent on the biological activity of a group of proteins collectively called interleukins that are secreted. Interleukins that have been studied in detail include interleukin-4 (IL-4). Two key molecules identified as ) and interleukin-13 (IL-13) It is included. IL-4 and IL-13 are CD4 + Th2 cells, natural killer T cells Cells are classified into numerous cell types, including NKT cells, macrophages, mast cells, and basophils. These are two closely related proteins that can be secreted. IL-4 and IL-13 are numerous. It exhibits overlapping functions and is critically important for the development of the T cell-dependent humoral immune response. Despite their similarities in formation, cell source, and biological function, these sites Each of the Cains mediates a specialized function, which is these interleukins The receptors and downstream signs mediate both their common and specific biological activities. This has stimulated numerous studies aimed at identifying naring pathways.

[0004] IL-4 has a high affinity for two receptors, namely type I and type II IL-4 receptors. It is currently known that it binds via Nity. The type I IL-4 receptor binds to the IL-4 receptor α chain. , several other interleukins including IL-2, IL-7, IL-9 and IL-15 It consists of a common γC chain, which is also part of the receptor. The type II IL-4 receptor is an IL-4 receptor. It consists of the somatic α chain and the IL-13 receptor α1 chain. On the other hand, IL-13 is a type II IL-4 receptor. It binds to the body and then to a specific receptor called the IL-13 receptor α2. IL-13 binding to receptor α2 does not transmit a signal, and this receptor also does not transmit signals in its soluble form. It is secreted. Therefore, the IL-13 receptor α2 is often referred to as a decoy receptor. .

[0005] Genes encoding the IL-4 protein have been cloned from various species, including bacteria and It is expressed in mammalian cells. For example, mature human IL-4 has an estimated 15Kd value. It is a secretory polypeptide consisting of 129 amino acids that has molecular weight, and it encodes human IL-4. cDNA indicates [Yokota et al., Proc Natl Acad Sci U SA.83(16):5894-5898(1986)]. Canine IL-4 protein The corresponding cDNA has also been identified, and it has 40% identity with human IL-4. It has been shown to encode a two-amino acid polypeptide [van der Kaai j et al., Immunogenetics 49:142-143 (1999). Human IL The gene encoding -13 has been cleaned and is expressed in various host systems. Minty et al., Nature 362:248-50 (1993). Mature IL-13 is It is a secreted polypeptide with an apparent molecular weight of 12.4 kD, and human IL-13 This is indicated by the cDNA encoding it. The cDNA encoding canine IL-13 has also been identified. [Yang et al., J. Interferon and Cytokine Research] [rch 20:779-785(2000)]. The estimated canine IL-13 mature polypeptide is It consists of 111 amino acids and has 61.8% identity with human IL-13.

[0006] Genes encoding the human and mouse IL-4 receptor α chain have been cloned, and various It is expressed in the host system. For example, the cDNA encoding the human IL-4 receptor α chain. Galizzi et al. [International Immunology 2(7): It is described in 669-675 (1990) and the mouse IL-4 receptor α chain is coated The cDNA used was identified by Mosley et al. [Cell, 59(2):335-348(1989)] It is described by [ ]. The cDNA of the human IL-4 receptor α chain is a 24-amino acid chain. It encodes 825 amino acid residues, including the Gunal sequence. The mouse protein is a human receptor. Although it is 15 amino acid residues shorter, both proteins are closely related and have identical overall sequences. Sex is 50% at the amino acid level.

[0007] Genes encoding the horse, canine, and feline IL-4 receptor α chain have also been disclosed [US See 7,208,579 B2. Also, the isoflavones of the canine IL-4 receptor α cDNA presumed to correspond to one of the forms has been found in the Genbank database. It can be released (SEQ ID NO: 1). Therefore, the present invention determines the IL-4 receptor α chain cDNA. We carried out the task of clearly determining the encoding polypeptide sequence.

[0008] IL-4 and IL-13 are extracellular pathogens (e.g., tissue or tubular parasites). It is a critically important cytokine for the development of the Th2 immune response necessary for defense against [the disease]. However, both cytokines are involved in various allergic diseases in humans and animals (asthma and It is associated with the pathogenesis of conditions including atopic dermatitis. Asthma is common in humans. It is a respiratory disease. This disease involves pneumonia, hypersensitivity of the bronchial airways to external stimuli, and trachea It is characterized by structural modifications of the musculoskeletal wall tissue. The pathophysiology of allergic asthma is described by Vatre. lla et al [Journal of Asthma and Allergy 7:123 This is outlined in -130 (2014). Asthma is caused by large amounts of IL-4 and IL-1 CD4 produces 3 and modulates the immune-inflammatory response in the allergic airways. + Th2 cells It is maintained. Recent advances in understanding the asthma response suggest that IL-4 is involved in the pathogenesis of the disease. It highlights the important roles played by both and IL-13. For example, both Cytokines also divide immunoglobulin isotypes in B cells, from IgM to IgE. By stimulating replacement, this allergen-specific IgE causes mast cell degranulation and inflammation in the airways. It contributes to the release of mediators. In addition, both IL-4 and IL-13 affect bronchial smooth muscle. This enhances the contraction of the eosinophils and stimulates airway recruitment of eosinophils, which in turn stimulates corresponding receptors on eosinophils. It can also induce degranulation in response to cross-linking of allergen-binding IgE to IL. -13 also stimulates mucus secretion and promotes airway remodeling, which is goblet cell hyperplasia, collagen It is caused by the deposition of enzymes and the proliferation of airway smooth muscle cells. Therefore, L-4 and IL-13 trigger the onset of asthma attacks, including bronchoconstriction and increased airway function. It is now clear that this is closely related to the pathological changes that lead to it.

[0009] Atopic dermatitis (AD) is characterized by dysregulation of the immune system and abnormalities of the epidermal barrier. It is a recurrent pruritic inflammatory skin disease. The pathological and immunological characteristics of AD have been extensively studied. [Rahman et al., Inflammation & Allergy] -drug target 10:486-496(2011) and Harskamp Seminar in Cutaneous Medicine and Surg This is outlined in ery 32:132-139 (2013). AD is the most common disease in humans. It is also a common skin disease, affecting 2-10% of the adult population in the United States and approximately 25% of children worldwide. % are affected. In humans, AD skin lesions involve Th2 cells, eosinophils, mast cells and dendritic cells. These lesions are characterized by cellular infiltration. In the acute phase of AD, these lesions are caused by IL-4 AD exhibits dominant expression of Th2-type cytokines, including IL-13. Also characterized by an increase in the level of skin CD4 + IL-4 in Th2 cells It is positively correlated with the expression of IL-13. AD is classified as a Th2 disease, but Th 1. Other T cell subsets, such as Th22 and Th17, also contribute to the pathogenesis of the disease. It is possible that... Despite the global increase in the incidence of Alzheimer's disease, the symptoms are... For patients whose condition is not adequately controlled by oral corticosteroids, the available treatment options include oral corticosteroids, and Oral cyclosporine and narrowband UVB phototherapy are the only treatments available. These therapies are not always effective. Moreover, their use comes with various safety implications. Recently, human IL-4R α Unique to Monoclonal antibodies have been developed, and some of these antibodies are used in the treatment of atopic dermatitis. Their therapeutic utility in humans has been extensively tested [e.g., US Please refer to 20150017176 A1.

[0010] Alzheimer's disease (AD) is a common disease in companion animals, especially dogs, and its prevalence in dogs is high. It is estimated that this accounts for approximately 10-15% of the total dog population. AD in dogs and cats The pathogenesis of [Nuttall et al., Veterinary Records 172(8)] [This is outlined in 201-207 (2013)] and shows a significant difference compared to the case of AD in humans. They share similarities, and these include CD4 with a dominance of IL-4 and IL-13 cytokines. -It includes a Th2-biased cytokine environment and infiltration of various immune cells into the skin. In humans Similar to the human case, current therapies for atopic dermatitis in dogs and cats are emollient therapies such as shampooing and moisturizers, or symptomatic therapies using oral or systemic corticosteroids and oral cyclosporine. Similar to the case of human AD, these therapies do not address the underlying mechanisms of the disease and have significant safety and efficacy issues. Therefore, there is an unmet medical need for safe and effective treatment options for AD in companion animals. Such treatment should preferably interfere with the underlying mechanisms of the disease.

[0011] The citation of any reference in this specification should not be construed as an admission that such reference is available as "prior art" for this application. SUMMARY OF THE INVENTION

[0012] SUMMARY OF THE INVENTION The present invention relates to an anti-canine interleukin α 4 receptor alpha (IL-4R ) antibody having high binding affinity for canine IL-4R α . In a more detailed embodiment the anti-canine interleukin 4 receptor alpha (IL-4R α ) antibody also has the ability to block the binding of canine IL-4 and canine IL-13 to type I or II IL-4 receptors and then suppress signaling from both canine IL -4 and IL-13. In certain embodiments such anti-canine IL-4R α antibodies are mouse anti-canine IL-4R α antibodies Yes. In a more detailed embodiment, the anti-canine IL-4R α The antibody is canine IL-4R α to It has high binding affinity to the canine IL-4 receptor, and to type I and type II IL-4 receptors. It also has the ability to block the binding of canine IL-13.

[0013] Furthermore, the present invention relates to the complementarity-determining regions (CDRs) contained in these antibodies, and these CDR (e.g., mouse anti-canine IL-4R) α (What is obtained from antibodies) inside the dog frame Incorporate canine-like anti-canine IL-4R α This invention relates to obtaining antibodies. The present invention also relates to atopic Conditions such as dermatitis, and / or type I and / or type II IL-4 receptors Due to the downstream signaling effects from the binding of canine IL-4 and / or canine IL-13 Regarding the use of such antibodies in the treatment of other harmful conditions.

[0014] Therefore, the present invention provides 14 illustrated mouse anti-canine IL-4R α C from antibodies It provides a specific set (combination) of DRs. 14 such exemplary mouse antibodies Inu IL-4R α The antibody is a specific set of CDRs, namely, three light chain CDRs called CD R light chain 1 (CDRL1), CDR light chain 2 (CDRL2), and CDR light chain 3 (CDRL3) ) and three heavy chain CDRs: CDR heavy chain 1 (CDRH1), CDR heavy chain 2 (CDR It has H2) and CDR heavy chain 3 (CDRH3). As described in detail below. There is considerable sequence homology and even some overlap within each group of CDRs (for example, later See the CDRL1 set described below. Therefore, the present invention is illustrated by 14 examples. Mouse anti-canine IL-4R α Simply provide the amino acid sequences of 6 CDRs from the antibody. Furthermore, there are also conservatively modified variants of these CDRs, as well as the same standard (Canonica (L) including structures (e.g., shared) and / or canine IL-4R α The epitope contains IL-4R α One or more amino acid residues (for example, 1 to 4 or more) It also provides mutants that bind to it.

[0015] Therefore, the present invention relates to Sequence ID No. 47, Sequence ID No. 48, Sequence ID No. 49, Sequence ID No. 50, Sequence ID 51, Sequence ID 52, Sequence ID 53, Sequence ID 54, Sequence ID 55, Sequence ID 1 29. Light chain complementarity determination region containing the amino acid sequence of SEQ ID NO: 130 or SEQ ID NO: 131 1 (VL CDR1) and / or SEQ ID NO: 56, SEQ ID NO: 57, SEQ ID NO: 581 Number 59, Sequence ID 60, Sequence ID 61, Sequence ID 62, Sequence ID 63, Sequence ID 64, Light chain complementation containing the amino acid sequence of SEQ ID NO: 132, SEQ ID NO: 133, or SEQ ID NO: 134 Sex determination region 2 (VL CDR2) and / or SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67, SEQ ID NO: 68, SEQ ID NO: 69, SEQ ID NO: 70, SEQ ID NO: 71, SEQ ID NO: 72, Array Number 73, Sequence ID 135, Sequence ID 136, Sequence ID 137, Sequence ID 138 or Light chain complementarity determination region 3 (VL CDR3) containing the amino acid sequence of SEQ ID NO: 139 and / Or sequence numbers 74, 75, 76, 77, 78, array Number 79, Sequence ID 80, Sequence ID 81, Sequence ID 82, Sequence ID 140, Sequence ID 14 1. Heavy chain complementarity determination region containing the amino acid sequence of SEQ ID NO: 142 or SEQ ID NO: 143 (VH CDR1) and / or Sequence ID 83, Sequence ID 84, Sequence ID 85, Sequence ID No. Number 86, Sequence ID 87, Sequence ID 88, Sequence ID 89, Sequence ID 90, Sequence ID 91, Distribution Column number 144, sequence number 145, sequence number 146, sequence number 147, or sequence number 14 Heavy chain complementarity determination region 2 (VH CDR2) and / or sequence number containing 8 amino acid sequences Number 92, Sequence ID 93, Sequence ID 94, Sequence ID 95, Sequence ID 96, Sequence ID 97, Distribution Column number 98, Sequence number 99, Sequence number 100, Sequence number 149, Sequence number 150, Sequence number Heavy chain complementarity determination including amino acid sequences of code 151, SEQ ID NO: 152, or SEQ ID NO: 153. IL-4R, including region 3 (VH CDR3) α Antibodies or antigens that specifically bind to them. It provides a binding fragment. In certain embodiments, the antibody is a mammalian antibody. In a more detailed embodiment, the antibody is a canine antibody.

[0016] Therefore, the canine antibody or its antigen-binding fragment of the present invention is SEQ ID NO: 74 , SEQ ID NO: 75, SEQ ID NO: 76, SEQ ID NO: 77, SEQ ID NO: 78, SEQ ID NO: 79, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 82, SEQ ID NO: 140, SEQ ID NO: 141, SEQ ID NO: 142 Or heavy chain complementarity determination region 1 (VH CDR1) having the amino acid sequence of SEQ ID NO: 143 It includes one or more of the following. In another embodiment, the heavy chain complementarity determination region 2 (VH CDR 2) Sequence numbers 83, 84, 85, 86, 87, sequence Number 88, Sequence ID 89, Sequence ID 90, Sequence ID 91, Sequence ID 144, Sequence ID 14 5. Contains the amino acid sequence of SEQ ID NO: 146, SEQ ID NO: 147, or SEQ ID NO: 148. In another embodiment, the heavy chain complementarity determination region 3 (VH CDR3) is SEQ ID NO: 9 2, Sequence ID 93, Sequence ID 94, Sequence ID 95, Sequence ID 96, Sequence ID 97, Sequence Number Number 98, Sequence ID 99, Sequence ID 100, Sequence ID 149, Sequence ID 150, Sequence ID 1 51, containing the amino acid sequence of SEQ ID NO: 152 or SEQ ID NO: 153. In the embodiment, the canine antibody or antigen-binding fragment is SEQ ID NO: 74, Column number 75, Sequence ID 76, Sequence ID 77, Sequence ID 78, Sequence ID 79, Sequence ID 80 , Sequence ID 81, Sequence ID 82, Sequence ID 140, Sequence ID 141, Sequence ID 142 also This includes VH CDR1 containing the amino acid sequence of SEQ ID NO: 143, and SEQ ID NO: 83, SEQ ID NO: 84 , SEQ ID NO: 85, SEQ ID NO: 86, SEQ ID NO: 87, SEQ ID NO: 88, SEQ ID NO: 89, SEQ ID NO: 90, SEQ ID NO: 91, SEQ ID NO: 144, SEQ ID NO: 145, SEQ ID NO: 146, SEQ ID NO: 14 It contains both VH CDR2 containing amino acid sequence 7 or sequence number 148. In such embodiments, the canine antibody or antigen-binding fragment is sequence Number 74, Sequence ID 75, Sequence ID 76, Sequence ID 77, Sequence ID 78, Sequence ID 79, Sequence ID 80, Sequence ID 81, Sequence ID 82, Sequence ID 140, Sequence ID 141, Sequence Number VH CDR1 containing the amino acid sequence of number 142 or SEQ ID NO: 143, and SEQ ID NO: 92, SEQ ID NOs: 93, 94, 95, 96, 97, 9 8, SEQ ID NO: 99 or SEQ ID NO: 100, SEQ ID NO: 149, SEQ ID NO: 150, SEQ ID NO: 1 51, and VH CDR3 containing the amino acid sequence of SEQ ID NO: 152 or SEQ ID NO: 153 This includes the method. In yet another such embodiment, the canine antibody or antigen binding. The sex fragments are sequence numbers 83, 84, 85, 86, and sequence number Number 87, Sequence ID 88, Sequence ID 89, Sequence ID 90, Sequence ID 91, Sequence ID 144, Amino acid sequences of SEQ ID NO: 145, SEQ ID NO: 146, SEQ ID NO: 147, or SEQ ID NO: 148 VH CDR2 including, and sequence numbers 92, 93, 94, 95, Sequence ID 96, Sequence ID 97, Sequence ID 98, Sequence ID 99 or Sequence ID 100, Array Number 149, Sequence ID 150, Sequence ID 151, Sequence ID 152 or Sequence ID 153 This includes both VH CDR3 containing an amino acid sequence. Yet another such embodiment In this case, the canine antibody or antigen-binding fragment is SEQ ID NO: 74, SEQ ID NO: 7 5, Sequence ID 76, Sequence ID 77, Sequence ID 78, Sequence ID 79, Sequence ID 80, Sequence No. Sequence No. 81, Sequence No. 82, Sequence No. 140, Sequence No. 141, Sequence No. 142 or Sequence No. VH CDR1 containing amino acid sequence 143, SEQ ID NO: 83, SEQ ID NO: 84, SEQ ID NO: 85, SEQ ID NO: 86, SEQ ID NO: 87, SEQ ID NO: 88, SEQ ID NO: 89, SEQ ID NO: 90, Array Number 91, Sequence ID 144, Sequence ID 145, Sequence ID 146, Sequence ID 147 or VH CDR2 containing the amino acid sequence of column number 148, and SEQ ID NOs: 92, SEQ ID NOs: 93 , SEQ ID NO: 94, SEQ ID NO: 95, SEQ ID NO: 96, SEQ ID NO: 97, SEQ ID NO: 98, SEQ ID NO: 99 or SEQ ID NO: 100, SEQ ID NO: 149, SEQ ID NO: 150, SEQ ID NO: 151, SEQ ID NO: 99, SEQ ID NO: 100, SEQ ID NO: 149, SEQ ID NO: 150, SEQ ID NO: 151, SEQ ID NO: 151 Contains VH CDR3 with the amino acid sequence of number 152 or SEQ ID NO: 153.

[0017] In certain embodiments, the canine antibody or antigen-binding fragment is also sequence Number 47, Sequence ID 48, Sequence ID 49, Sequence ID 50, Sequence ID 51, Sequence ID 52, Sequence IDs 53, 54, 55, 129, 130 or It contains light chain complementarity determination region 1 (VL CDR1), which includes the amino acid sequence at column number 131. In the linked embodiment, the light chain complementarity determination region 2 (VL CDR2) is sequence number 56, sequence number Number 57, Sequence ID 58, Sequence ID 59, Sequence ID 60, Sequence ID 61, Sequence ID 62, Sequence ID 63, Sequence ID 64, Sequence ID 132, Sequence ID 133 or Sequence ID 134 It includes an amino acid sequence. In yet another embodiment, the light chain complementarity determination region 3(VL) CDR3) contains sequence numbers 65, 66, 67, 68, and 6 9, SEQ ID NO: 70, SEQ ID NO: 71, SEQ ID NO: 72, SEQ ID NO: 73, SEQ ID NO: 135, Array Contains the amino acid sequence of number 136, SEQ ID NO: 137, SEQ ID NO: 138, or SEQ ID NO: 139 In this particular embodiment, the canine antibody or antigen-binding fragment The sequence is sequence number 47, sequence number 48, sequence number 49, sequence number 50, sequence number 51, sequence Number 52, Sequence ID 53, Sequence ID 54, Sequence ID 55, Sequence ID 129, Sequence ID 13 VL CDR1 containing amino acid sequence 0 or sequence number 131, and sequence number 56, sequence number Number 57, Sequence ID 58, Sequence ID 59, Sequence ID 60, Sequence ID 61, Sequence ID 62, Distribution Column number 63, sequence number 64, sequence number 132, sequence number 133 or sequence number 134 It includes both VL CDR2 containing the minoic acid sequence.

[0018] In other such embodiments, the canine antibody or antigen-binding fragment is Sequence ID 47, Sequence ID 48, Sequence ID 49, Sequence ID 50, Sequence ID 51, Sequence ID 5 2. Sequence ID 53, Sequence ID 54, Sequence ID 55, Sequence ID 129, Sequence ID 130, This includes VL CDR1 containing the amino acid sequence of SEQ ID NO: 131, and SEQ ID NO: 65, SEQ ID NO: 66 , SEQ ID NO: 67, SEQ ID NO: 68, SEQ ID NO: 69, SEQ ID NO: 70, SEQ ID NO: 71, SEQ ID NO: 72, SEQ ID NO: 73, SEQ ID NO: 135, SEQ ID NO: 136, SEQ ID NO: 137, SEQ ID NO: 13 It contains both VL CDR3 containing amino acid sequence 8 or sequence number 139. In one such embodiment, the canine antibody or antigen-binding fragment is Sequence ID 56, Sequence ID 57, Sequence ID 58, Sequence ID 59, Sequence ID 60, Sequence ID 6 1. Sequence ID 62, Sequence ID 63, Sequence ID 64, Sequence ID 132, Sequence ID 133, This includes VL CDR2 containing the amino acid sequence of SEQ ID NO: 134, and SEQ ID NO: 65, SEQ ID NO: 66 , SEQ ID NO: 67, SEQ ID NO: 68, SEQ ID NO: 69, SEQ ID NO: 70, SEQ ID NO: 71, SEQ ID NO: 72, SEQ ID NO: 73, SEQ ID NO: 135, SEQ ID NO: 136, SEQ ID NO: 137, SEQ ID NO: 13 It contains both VL CDR3 containing amino acid sequence 8 or sequence number 139.

[0019] In yet other such embodiments, the canine antibody or antigen-binding fragment This includes sequence numbers 47, 48, 49, 50, 51, and sequence number Sequence ID 52, Sequence ID 53, Sequence ID 54, Sequence ID 55, Sequence ID 129, Sequence ID 130 Alternatively, VL CDR1 containing the amino acid sequence of SEQ ID NO: 131, SEQ ID NO: 56, SEQ ID NO: 5 7, Sequence ID 58, Sequence ID 59, Sequence ID 60, Sequence ID 61, Sequence ID 62, Sequence Number Amino acid of number 63, SEQ ID NO: 64, SEQ ID NO: 132, SEQ ID NO: 133, or SEQ ID NO: 134 VL CDR2 containing acid sequences, and SEQ ID NOs. 65, 66, 67, sequence Number 68, Sequence ID 69, Sequence ID 70, Sequence ID 71, Sequence ID 72, Sequence ID 73, SEQ ID NO: 135, SEQ ID NO: 136, SEQ ID NO: 137, SEQ ID NO: 138 or SEQ ID NO: 13 Contains VL CDR3 with a 9-amino acid sequence.

[0020] In certain embodiments, the canine anti-canine IL-4R α Antibodies have a complementarity-determining region (CD). R) is included, in this case CDR is with respect to the heavy chain CDR1, CDR2 and CDR3. Each has the standard (canonical) structure of H1-1, H2-3A, and H3-12, i.e. The heavy chain CDR1 has standard structural class 1, and the heavy chain CDR2 has standard structural class 3A. Furthermore, the heavy chain CDR3 has standard structural class 12. In a more detailed embodiment, The corresponding CDRs for the light chains are L1- for CDR1, CDR2, and CDR3 of the light chains. 1, has the standard structure of L2-1 and L3-1. In other embodiments, the canine anti Inu IL-4R α Antibodies contain a complementation-determining region (CDR), in which case the CDR is the C of the heavy chain. H1-1, H2-2A, and H3-7 respectively apply to DR1, CDR2, and CDR3. It has a standard structure. In a more detailed embodiment of this type, the CD of the corresponding light chain R is L1-2A and L2-1 for the light chains CDR1, CDR2, and CDR3, respectively. It has the standard structure of L3-1. In yet another embodiment, the canine anti-canine IL- 4R α Antibodies contain a complementation-determining region (CDR), in this case the CDR is CDR1 of the heavy chain, C Standard structures for DR2 and CDR3 are H1-1, H2-2B, and H3-15, respectively. It has a structure. In a more detailed embodiment of this type, the CDR of the corresponding light chain is the light chain Regarding CDR1, CDR2, and CDR3, L1-4, L2-1, and L3- respectively It has a standard structure of 1. In yet another embodiment, the canine anti-canine IL-4R α antibody It includes a complementarity-determining region (CDR), in which case the CDR is CDR1, CDR2 and Regarding CDR3, it has standard structures H1-1, H2-1, and H3-15, respectively. In a more detailed embodiment of this type, the CDR of the corresponding light chain is the CDR1 of the light chain. Standard structures for CDR2 and CDR3: L1-3, L2-1, and L3-1, respectively. It has. In yet another embodiment, the canine anti-canine IL-4R α Antibodies determine complementarity. Includes a region (CDR), in which case the CDR is CDR1, CDR2, and CDR3 of the heavy chain. Regarding this, there are standard structures of H1-1, H2-2B, and H3-6, respectively. In a more detailed embodiment, the CDR of the corresponding light chain is CDR1, CDR2 and The standard structures for both and CDR3 are L1-2A, L2-1, and L3-1, respectively.

[0021] In yet another embodiment, the canine anti-canine IL-4R α Antibodies are found in the complementarity-determining region (C (DR) is included, in this case CDR is with respect to the heavy chain CDR1, CDR2 and CDR3 Each has a standard structure of H1-1, H2-2B, and H3-4. In detailed embodiments, the CDRs of the corresponding light chains are CDR1, CDR2 and CDR Regarding 3, there are standard structures L1-6, L2-1, and L3-1, respectively. Furthermore, other practical In terms of application method, the canine anti-canine IL-4R αAntibodies contain a complementation-determining region (CDR). In this case, CDR is H1- with respect to the heavy chains CDR1, CDR2, and CDR3, respectively. 1. It has the standard structure of H2-1 and H3-13. A more detailed implementation of this type In this state, the CDR of the corresponding light chain is related to the CDR1, CDR2 and CDR3 of the light chain. Each has a standard structure of L1-1, L2-1, and L3-1, respectively. Furthermore, in other embodiments... Therefore, the canine anti-canine IL-4R α Antibodies contain a complementation-determining region (CDR), in this case, CDR is H1-1, H2-2, and H2-2 respectively for the heavy chain CDR1, CDR2, and CDR3. It has the standard structure of A and H3-6. In a more detailed embodiment of this type, The corresponding CDRs for the light chains are L1- for CDR1, CDR2, and CDR3 of the light chains. It has standard structures of 2A, L2-1, and L3-1.

[0022] In yet another embodiment, the canine anti-canine IL-4R α Antibodies are found in the complementarity-determining region (C (DR) is included, in this case CDR is with respect to the heavy chain CDR1, CDR2 and CDR3 Each has a standard structure of H1-1, H2-3A, and H3-15 or H3-13, respectively. In a more detailed embodiment of this type, the CDR of the corresponding light chain is the CDR1 of the light chain, C The standard structures for DR2 and CDR3 are L1-6, L2-1, and L3-1, respectively. It possesses. In yet another embodiment, the canine anti-canine IL-4R α Antibodies are the complementarity-determining region. Includes a region (CDR), in which case the CDR is related to the heavy chain CDR1, CDR2, and CDR3. These have standard structures of H1-1, H2-2A, and H3-10, respectively. In a more detailed embodiment, the CDR of the corresponding light chain is CDR1, CDR2 and Regarding CDR3, it has standard structures L1-6, L2-1, and L3-1, respectively. In other embodiments, the canine anti-canine IL-4R α Antibodies are found in the complementarity-determining region (CDR). ) including, in this case CDR is CDR1, CDR2 and CDR3 of the heavy chain respectively It has standard structures of H1-1, H2-3A, and H3-9. Further details of this type In one embodiment, the CDR of the corresponding light chain is CDR1, CDR2 and CDR3 of the light chain. Regarding these, they have standard structures L1-3, L2-1, and L3-3, respectively.

[0023] The present invention also relates to IL-4R, which comprises a canine IgG heavy chain and a canine kappa or lambda light chain. α Provides isolated canine antibodies or their antigen-binding fragments that specifically bind to a particular antigen. In this particular embodiment, there are three light chain complementarity determining regions (CDRs). CDR light chain 1 (CDRL1), CDR light chain 2 (CDRL2), and CDR light chain 3 (CDR Inukappa or lambda light chains including L3), and CDR heavy chains which are three heavy chain CDRs. Contains 1 (CDRH1), CDR heavy chain 2 (CDRH2), and CDR heavy chain 3 (CDRH3). The canine IgG heavy chain is a mouse anti-canine IL-4R α Obtained from antibodies. The canine antibody of the present invention And a specific embodiment of its antigen-binding fragment is canine IL-4R α Combined, and / or Canine IL-4 to Canine IL-4R α It blocks the bond.

[0024] In certain embodiments, the present invention relates to three light chain complementarity determination regions (CDRs) C DR light chain 1 (CDRL1), CDR light chain 2 (CDRL2), and CDR light chain 3 (CDRL 3) and three heavy chain CDRs: CDR heavy chain 1 (CDRH1), CDR heavy chain 2 (CDRH 2) and CDR heavy chain 3 (CDRH3), canine interleukin 4 receptor alpha (IL-4R α Isolated mammalian antibodies that specifically bind to ) or their antigen-binding fura Provides a component. In one embodiment, CDRL1 is sequence number 47, sequence number 4 7 variants, conserved modified variant of sequence number 47 (i.e., a conserved modified variant) , amino acid sequence of the variant of SEQ ID NO: 47 containing 1 standard structural class, SEQ ID NO: 48, sequence Sequences containing mutant number 48, a conserved modified mutant of sequence number 48, and the standard structural class of 2A. A variant of number 48, sequence number 49, a variant of sequence number 49, a conservative modification variant of sequence number 49. Variants of SEQ ID NO: 49, including four standard structural classes, SEQ ID NO: 50, SEQ ID NO: 50 Mutants, Conservative Modified Mutants of SEQ ID NO: 50, Mutants of SEQ ID NO: 50 Including 3 Standard Structure Classes Allomorph, SEQ ID NO: 51, mutant of SEQ ID NO: 51, conservative modified mutant of SEQ ID NO: 51, 3 labels Variants of SEQ ID NO: 51 containing substructure classes, SEQ ID NO: 52, variants of SEQ ID NO: 52, sequence Conservative modified mutant of number 52, mutant of sequence number 52 containing the standard structural class 2A, sequence Number 53, mutant of SEQ ID NO: 53, conservative modified mutant of SEQ ID NO: 53, standard structural class 6 A variant of SEQ ID NO: 53 containing SEQ ID NO: 54, a variant of SEQ ID NO: 54, and SEQ ID NO: 54 Conservative modified mutant, mutant of SEQ ID NO: 54 containing one standard structural class, SEQ ID NO: 55, The variant of sequence number 55, the conserved modified variant of sequence number 55, and the standard structural class of 2A are included in the distribution. Variant of sequence number 55, sequence number 129, variant of sequence number 129, storage of sequence number 129 Modified mutants, mutant of SEQ ID NO: 129 containing 6 standard structural classes, SEQ ID NO: 130, Includes the mutant with sequence number 130, the conserved modified mutant with sequence number 130, and 6 standard structure classes. Variant of SEQ ID NO: 130, SEQ ID NO: 131, Variant of SEQ ID NO: 131 The amino acid sequences of the conserved modified mutant, the mutant of SEQ ID NO: 131, which includes three standard structural classes. include.

[0025] The corresponding CDRL2s are sequence number 56, a variant of sequence number 56, and a conservative modified version of sequence number 56. Modified mutant, mutant of SEQ ID NO: 56 containing one standard structural class, SEQ ID NO: 57, SEQ ID NO: 5 7 variants, a conserved modified variant of SEQ ID NO: 57, and SEQ ID NO: 57 including one standard structure class. mutant of, SEQ ID NO: 58, mutant of SEQ ID NO: 58, conservative modified mutant of SEQ ID NO: 58, 1 The variant of SEQ ID NO: 58, SEQ ID NO: 59, and the variant of SEQ ID NO: 59, which include the standard structural class. Conserved modified mutant of SEQ ID NO: 59, mutant of SEQ ID NO: 59 containing 1 standard structure class, Column number 60, mutant of sequence number 60, conservative modified mutant of sequence number 60, standard structure of 1 Variants of SEQ ID NO: 60 including Ras, SEQ ID NO: 61, variants of SEQ ID NO: 61, SEQ ID NO: 61 Conserved modified mutants of, mutant of SEQ ID NO: 61 containing 1 standard structure class, SEQ ID NO: 62, A variant of SEQ ID NO: 62, a conserved modified variant of SEQ ID NO: 62, and a standard structural class containing 1. Variant of sequence number 62, SEQ ID NO: 63, Variant of SEQ ID NO: 63, Conservative modification of SEQ ID NO: 63 Mutant, mutant of SEQ ID NO: 63 containing standard structural class 1, SEQ ID NO: 64, SEQ ID NO: 64 A variant of, a conserved modified variant of SEQ ID NO: 64, and a standard structural class of SEQ ID NO: 64 Mutant, SEQ ID NO: 132, mutant of SEQ ID NO: 132, conservative modified mutant of SEQ ID NO: 132 , a variant of SEQ ID NO: 132 containing a standard structural class, SEQ ID NO: 133, SEQ ID NO: 133 A variant of, a conserved modified variant of SEQ ID NO: 133, and SEQ ID NO: 133 containing a standard structure class of 1. 3 variants, SEQ ID NO: 134, SEQ ID NO: 134 variant, SEQ ID NO: 134 conservative modification variant Includes the amino acid sequence of a variant of SEQ ID NO: 134, which is an isomorph or contains one standard structural class.

[0026] The corresponding CDRL3s are sequence number 65, a variant of sequence number 65, and a conservative modified version of sequence number 65. Decorative mutants, mutant of SEQ ID NO: 65 containing one standard structural class, SEQ ID NO: 66, SEQ ID NO: 6 6 variants, a conserved modified variant of SEQ ID NO: 66, and SEQ ID NO: 66 including one standard structure class. mutant of, SEQ ID NO: 67, mutant of SEQ ID NO: 67, conservative modified mutant of SEQ ID NO: 67, 1 The variant of SEQ ID NO: 67, SEQ ID NO: 68, and the variant of SEQ ID NO: 68, which include the standard structural class. Conservative modified mutant of SEQ ID NO: 68, mutant of SEQ ID NO: 68 containing 1 standard structure class, Column number 69, mutant of sequence number 69, conservative modified mutant of sequence number 69, standard structure of 1 Variants of SEQ ID NO: 69 including Ras, SEQ ID NO: 70, variants of SEQ ID NO: 70, SEQ ID NO: 70 Conserved modified variants of, variant of SEQ ID NO: 70 containing 1 standard structure class, SEQ ID NO: 71, A variant of SEQ ID NO: 71, a conserved modified variant of SEQ ID NO: 71, and a standard structural class of 1. Variants of sequence number 71, sequence number 72, variants of sequence number 72, and conservative modifications of sequence number 72. Mutant, mutant of SEQ ID NO: 72 containing standard structural class 1, SEQ ID NO: 73, SEQ ID NO: 73 A variant of SEQ ID NO: 73, a conserved modified variant of SEQ ID NO: 73, and a standard structural class of SEQ ID NO: 73 Mutant, SEQ ID NO: 135, mutant of SEQ ID NO: 135, conservative modified mutant of SEQ ID NO: 135 , variant of SEQ ID NO: 135, SEQ ID NO: 136, SEQ ID NO: 136, containing 1 standard structural class A variant of , a conserved modified variant of SEQ ID NO: 136, and SEQ ID NO: 13 containing a standard structure class of 1. 6 variants, SEQ ID NO: 137, SEQ ID NO: 137 variant, SEQ ID NO: 137 conservative modification variant Variants of SEQ ID NO: 137, SEQ ID NO: 138, SEQ ID NO: 1, including a variant of SEQ ID NO: 1, a standard structural class of SEQ ID NO: 1 38 variants, the conserved modified variant of SEQ ID NO: 138, and SEQ ID NO: 3 standard structure classes 138 variants, SEQ ID NO: 139, variant of SEQ ID NO: 139, conservative modification of SEQ ID NO: 139 Includes the amino acid sequence of the variant of SEQ ID NO: 139, which is either a decorative variant or a standard structural class of 1. .

[0027] The corresponding CDRH1 is sequence number 74, a variant of sequence number 74, and a conservative modified version of sequence number 74. Modified variants, variant of SEQ ID NO: 74 containing one standard structural class, SEQ ID NO: 75, SEQ ID NO: 7 5 variants, a conserved modified variant of SEQ ID NO: 75, and SEQ ID NO: 75 including one standard structure class. variants of, SEQ ID NO: 76, variants of SEQ ID NO: 76, and conservative modified variants of SEQ ID NO: 76. This includes a variant of SEQ ID NO: 76 containing one standard structural class, SEQ ID NO: 77, and a variant of SEQ ID NO: 77. The body, a conserved modified variant of SEQ ID NO: 77, or a variant of SEQ ID NO: 77 containing one standard structural class. Allomorph, SEQ ID NO: 78, mutant of SEQ ID NO: 78, conservative modified mutant of SEQ ID NO: 78, label 1 Variants of SEQ ID NO: 78, SEQ ID NO: 79, variants of SEQ ID NO: 79, and sequences containing substructure classes. Conservative modified variant of sequence number 79, variant of sequence number 79 containing 1 standard structure class, sequence number Syntax 80, mutant of SEQ ID NO: 80, conservative modified mutant of SEQ ID NO: 80, standard structural class of 1. Variants of SEQ ID NO: 80, SEQ ID NO: 81, variants of SEQ ID NO: 81, and preserved SEQ ID NO: 81 Existentially modified mutant, mutant of SEQ ID NO: 81 containing 1 standard structure class, SEQ ID NO: 82, sequence A variant of number 82, a conserved modified variant of sequence number 82, or a standard structural class of 1. Variant of sequence number 82, SEQ ID NO: 140, Variant of SEQ ID NO: 140, Storage of SEQ ID NO: 140 Modified mutants, mutant of SEQ ID NO: 140 containing 1 standard structural class, SEQ ID NO: 141, Includes the mutant of sequence number 141, the conserved modified mutant of sequence number 141, and the standard structure class of 1. Variants of SEQ ID NO: 141, SEQ ID NO: 142, Variants of SEQ ID NO: 142 Conservative modified mutant, mutant of SEQ ID NO: 142 containing standard structural class 1, SEQ ID NO: 143 , a variant of SEQ ID NO: 143, a conserved modified variant of SEQ ID NO: 143, or a standard structural variant of 1 Includes the amino acid sequence of the variant of SEQ ID NO: 143 containing the element S.

[0028] The corresponding CDRH2 is sequence number 83, a variant of sequence number 83, and a conservative modified version of sequence number 83. Modified mutants, variants of SEQ ID NO: 83 including the standard structural class 3A, SEQ ID NO: 84, SEQ ID NO: SEQ ID NOs: 84 variants, a conserved modified variant of SEQ ID NO: 84, and a standard structural class of SEQ ID NOs: 2A. 84 variants, SEQ ID NO: 85, SEQ ID NO: 85 variants, SEQ ID NO: 85 conservative modified variants Alternatively, variants of SEQ ID NO: 85, SEQ ID NO: 86, and SEQ ID NO: 86, which include the standard structural class 2B. mutants of, conserved modified mutant of SEQ ID NO: 86, SEQ ID NO: 87, mutant of SEQ ID NO: 87, Conservative modified mutant of sequence number 87, mutant of sequence number 87 containing 1 standard structure class, sequence Number 88, mutant of SEQ ID NO: 88, conservative modified mutant of SEQ ID NO: 88, standard structure of 2B Variants of SEQ ID NO: 88 including Ras, SEQ ID NO: 89, variants of SEQ ID NO: 89, SEQ ID NO: 89 Conservative modified mutants of 2B, mutant of SEQ ID NO: 89 including the standard structural class of 2B, SEQ ID NO: 90 This includes a variant of SEQ ID NO: 90, a conserved modified variant of SEQ ID NO: 90, and a standard structural class of 1. Variants of SEQ ID NO: 90, SEQ ID NO: 91, variants of SEQ ID NO: 91, conservative modifications of SEQ ID NO: 91 Decorative mutant, mutant of SEQ ID NO: 91 including the standard structural class 2A, SEQ ID NO: 144, sequence number A variant of number 144, a conserved modified variant of sequence number 144, and a standard structural class of 3A are included. Variant of sequence number 144, SEQ ID NO: 145, Variant of SEQ ID NO: 145, Conservative of SEQ ID NO: 145 Existentially modified mutants, mutants of SEQ ID NO: 145 including the standard structural class of 2A, SEQ ID NO: 146 , mutant of SEQ ID NO: 146, conservative modified mutant of SEQ ID NO: 146, standard structural class of 3A Variants of SEQ ID NO: 146, SEQ ID NO: 147, variants of SEQ ID NO: 147, SEQ ID NO: 1 47 conserved modified mutants, a mutant of sequence number 147 including the standard structural class of 3A, sequence number Label 148, variant of SEQ ID NO: 148, conservative modified variant of SEQ ID NO: 148, or label 3A This includes the amino acid sequence of a variant of SEQ ID NO: 148 that contains a substructure class.

[0029] The corresponding CDRH3 is sequence number 92, a variant of sequence number 92, and a conservative modified version of sequence number 92. Decorative mutants, mutants of SEQ ID NO: 92 including 12 standard structural classes, SEQ ID NO: 93, SEQ ID NO: SEQ ID NO: 93 includes 93 variants, a conserved modified variant of SEQ ID NO: 93, and 7 standard structural classes. 3 variants, SEQ ID NO: 94, SEQ ID NO: 94 variant, SEQ ID NO: 94 conservative modified variant or variants of SEQ ID NO: 94, SEQ ID NO: 95, SEQ ID NO: 95, which include 15 standard structural classes. Variants, including the conserved modified variant of SEQ ID NO: 95 or 11 standard structural classes of SEQ ID NO: 95 5 variants, SEQ ID NO: 96, SEQ ID NO: 96 variants, SEQ ID NO: 96 conservative modified variants, Variants of SEQ ID NO: 96, SEQ ID NO: 97, and SEQ ID NO: 97, which include 15 standard structural classes. The body, a conserved modified mutant of SEQ ID NO: 97, and a mutant of SEQ ID NO: 97 containing 6 standard structural classes. , SEQ ID NO: 98, mutant of SEQ ID NO: 98, conservative modified mutant of SEQ ID NO: 98, 4 standard constructs Variants of SEQ ID NO: 98, SEQ ID NO: 99, variants of SEQ ID NO: 99, SEQ ID NO: 99 conserved modified mutants, mutants of SEQ ID NO: 99 including 13 standard structure classes, SEQ ID NO: 100, a variant of sequence number 100, a conservative modified variant of sequence number 100, or a standard variant of 6 Variants of SEQ ID NO: 100, SEQ ID NO: 149, variants of SEQ ID NO: 149, and distribution Conservative modified mutant of sequence number 149, mutant of sequence number 149 containing 15 standard structure classes , SEQ ID NO: 150, mutant of SEQ ID NO: 150, conservative modified mutant of SEQ ID NO: 150, 10 Variants of SEQ ID NO: 150, SEQ ID NO: 151, and SEQ ID NO: 151, which include the standard structural class of SEQ ID NO: 150. Allomorphs, conserved modified variants of SEQ ID NO: 151, and SEQ ID NO: 151 including 15 standard structural classes A variant of, SEQ ID NO: 152, a variant of SEQ ID NO: 152, a conservative modification variant of SEQ ID NO: 152 The body, the variant of SEQ ID NO: 152 containing 9 standard structural classes, SEQ ID NO: 153, SEQ ID NO: 15 Sequences containing 3 variants, the conserved modified variant of sequence number 153, or 13 standard structural classes. Includes the amino acid sequence of mutant number 153.

[0030] In certain embodiments, the mammalian antibody of the present invention (including a chimeric mammalian antibody) and / Alternatively, the antigen-binding fragment is the canine interleukin-4 receptor alpha (IL-4). R α ) binds to and / or to canine IL-4 and / or canine IL-13 NuIL-4R α It blocks the binding of the mammalian antibody of the present invention and / or its antigen-binding fragment is the IL-4 type I receptor and / or IL-4 Blocks the binding of canine IL-4 and / or canine IL-13 to type II receptors. In the embodiment, the mammalian antibody (regardless of whether it is isolated or not) is a canine antibody. ru.

[0031] Therefore, in some embodiments of mammalian antibodies (including canine antibodies), CDRL1 This refers to SEQ ID NO: 47, a variant of SEQ ID NO: 47, a conservative modified variant of SEQ ID NO: 47, or a variant of SEQ ID NO: 1. Includes amino acid sequences of variants of SEQ ID NO: 47, which include substructure classes; CDRL2 is SEQ ID NO: 56, a variant of SEQ ID NO: 56, a conserved modified variant of SEQ ID NO: 56, or a standard structural variant of 1. Includes the amino acid sequence of the variant in SEQ ID NO: 56; CDRL3 is SEQ ID NO: 65, sequence A variant of number 65, a conserved modified variant of sequence number 65, or a standard structural class of 1. Includes the amino acid sequence of the variant in column number 65; CDRH1 is sequence number 74, sequence number 74 A variant, a conserved modified variant of SEQ ID NO: 74, or SEQ ID NO: 74 containing one standard structural class. Includes the amino acid sequence of the variant; CDRH2 is a variant of SEQ ID NO: 83, and is a variant of SEQ ID NO: 83. The conserved modified mutant of sequence number 83 and the mutant of sequence number 83 containing the standard structural class of 3A. The amino acid sequence includes; CDRH3 is a variant of SEQ ID NO: 92, SEQ ID NO: 92 The amino of the mutant of SEQ ID NO: 92, which includes 2 conserved modified mutants or 12 standard structural classes. Contains acid sequences.

[0032] In still other embodiments of mammalian antibodies (including canine antibodies), CDRL1 comprises the amino acid sequence of SEQ ID NO: 48, a variant of SEQ ID NO: 48, a conservatively modified variant of SEQ ID NO: 48, or a variant of SEQ ID NO: 48 that includes the standard structure class of 2A; CDRL2 comprises the amino acid sequence of SEQ ID NO: 57, a variant of SEQ ID NO: 57, a conservatively modified variant of SEQ ID NO: 57, or a variant of SEQ ID NO: 57 that includes the standard structure class of 1; CDRL3 comprises the amino acid sequence of SEQ ID NO: 66, a variant of SEQ ID NO: 66, a conservatively modified variant of SEQ ID NO: 66, or a variant of SEQ ID NO: 66 that includes the standard structure class of 1; CDRH1 comprises the amino acid sequence of SEQ ID NO: 75, a variant of SEQ ID NO: 75, a conservatively modified variant of SEQ ID NO: 75, or a variant of SEQ ID NO: 75 that includes the standard structure class of 1; CDRH2 comprises the amino acid sequence of SEQ ID NO: 84, a variant of SEQ ID NO: 84, a conservatively modified variant of SEQ ID NO: 84, and a variant of SEQ ID NO: 84 that includes the standard structure class of 2A; CDRH3 comprises the amino acid sequence of SEQ ID NO: 93, a variant of SEQ ID NO: 93, a conservatively modified variant of SEQ ID NO: 93, or a variant of SEQ ID NO: 93 that includes the standard structure class of 7. In still other embodiments of mammalian antibodies (including canine antibodies), CDRL1 comprises the amino acid sequence of SEQ ID NO: 49, a variant of SEQ ID NO: 49, a conservatively modified variant of SEQ ID NO: 49, or a variant of SEQ ID NO: 49 that includes the standard structure class of 4; CDRL2 comprises the amino acid sequence of SEQ ID NO: 58, a variant of SEQ ID NO: 58, a conservatively modified variant of SEQ ID NO: 58, or a variant of SEQ ID NO: 58 that includes the standard structure class of 4; CDRL3 comprises the amino acid sequence of SEQ ID NO: 67, a variant of SEQ ID NO: 67, a conservatively modified variant of SEQ ID NO: 67, or a variant of SEQ ID NO: 67 that includes the standard structure class of 4; CDRH1 comprises the amino acid sequence of SEQ ID NO: 76, a variant of SEQ ID NO: 76, a conservatively modified variant of SEQ ID NO: 76, or a variant of SEQ ID NO: 76 that includes the standard structure class of 4; CDRH2 comprises the amino acid sequence of SEQ ID NO: 85, a variant of SEQ ID NO: 85, a conservatively modified variant of SEQ ID NO: 85, or a variant of SEQ ID NO: 85 that includes the standard structure class of 4;

[0033] CDRH3 comprises the amino acid sequence of SEQ ID NO: 94, a variant of SEQ ID NO: 94, a conservatively modified variant of SEQ ID NO: 94, or a variant of SEQ ID NO: 94 that includes the standard structure class of 7. A variant of, a conserved modified variant of SEQ ID NO: 67, or a standard structural class of SEQ ID NO: 6 Includes the amino acid sequence of 7 variants; CDRH1 is the variant of SEQ ID NO: 76, SEQ ID NO: 76, Conservative modified mutants of SEQ ID NO: 76 or mutants of SEQ ID NO: 76 containing one standard structure class. The amino acid sequence includes; CDRH2 is a variant of SEQ ID NO: 85, SEQ ID NO: 8 The amino acids of the mutant of SEQ ID NO: 85, including the 5-conserved modified mutant and the 2B standard structure class. Contains acid sequence; CDRH3 is sequence number 94, a variant of sequence number 94, a conserved sequence number 94. The amino acid sequences of modified mutants or mutants of SEQ ID NO: 94 that include 15 standard structural classes include.

[0034] In yet another embodiment of mammalian antibodies (including canine antibodies), CDRL1 is sequence number Sequence number 51, a variant of SEQ ID NO: 51, a conserved modified variant of SEQ ID NO: 51, or a standard structure variant of SEQ ID NO: 3. Includes the amino acid sequence of the variant of SEQ ID NO: 51 containing Ras; CDRL2 is SEQ ID NO: 60, Includes variants of sequence number 60, conserved modified variants of sequence number 60, or standard structure class 1. Includes the amino acid sequence of the variant of SEQ ID NO: 60; CDRL3 is SEQ ID NO: 69, SEQ ID NO: 69 A variant of, a conserved modified variant of SEQ ID NO: 69, or a standard structural class of SEQ ID NO: 6 Includes the amino acid sequences of 9 variants; CDRH1 is a variant of SEQ ID NO: 78, SEQ ID NO: 78, Conservative modified mutants of SEQ ID NO: 78 or mutants of SEQ ID NO: 78 containing one standard structure class. The amino acid sequence includes; CDRH2 is a variant of SEQ ID NO: 87, SEQ ID NO: 8 Amino acids of the mutant of SEQ ID NO: 87, including 7 conserved modified mutants and 1 standard structural class. Includes sequence; CDRH3 is sequence number 96, variant of sequence number 96, conserved sequence number 96 Includes the amino acid sequence of the modified mutant or the mutant of SEQ ID NO: 96, which includes 15 standard structural classes. nothing.

[0035] In yet another embodiment of mammalian antibodies (including canine antibodies), CDRL1 is sequence number Syntax 52, mutant of SEQ ID NO. 52, conservative modified mutant of SEQ ID NO. 52, or standard structure of 2A Includes the amino acid sequence of the variant in SEQ ID NO: 52, which includes the class; CDRL2 is in SEQ ID NO: 61. Includes variants of SEQ ID NO: 61, conserved modified variants of SEQ ID NO: 61, or standard structural classes of 1. Includes the amino acid sequence of the variant of SEQ ID NO: 61; CDRL3 is SEQ ID NO: 70, SEQ ID NO: 7 SEQ ID NOs: 0 variants, SEQ ID NOs: 70 conservative modified variants, or SEQ ID NOs: 1 standard structure class Includes amino acid sequences of 70 variants; CDRH1 is a variant of SEQ ID NO: 79, SEQ ID NO: 79 , a conserved modified variant of SEQ ID NO: 79 or a variant of SEQ ID NO: 79 containing a standard structural class 1 Includes the amino acid sequence of the body; CDRH2 is a variant of SEQ ID NO: 88, SEQ ID NO: 88, SEQ ID NO: The mutant of SEQ ID NO: 88, including 88 conserved modified mutants and the standard structure class of 2B. Contains a noacid sequence; CDRH3 is a variant of SEQ ID NO 97, a variant of SEQ ID NO 97, a retainer of SEQ ID NO 97 Amino acid sequences of spatially modified mutants or mutants of SEQ ID NO: 97 that include 16 standard structural classes. Includes.

[0036] In yet another embodiment of mammalian antibodies (including canine antibodies), CDRL1 is sequence number Syntax 53, a variant of SEQ ID NO. 53, a conserved modified variant of SEQ ID NO. 53, or a standard structural variant of SEQ ID NO. 6. Includes the amino acid sequence of the variant of SEQ ID NO: 53 containing Ras; CDRL2 is SEQ ID NO: 62, Includes variants of sequence number 62, conserved modified variants of sequence number 62, or standard structural class 1. comprising the amino acid sequence of the variant of SEQ ID NO: 62; CDRL3 comprises SEQ ID NO: 71, the variant of SEQ ID NO: 71 a variant of SEQ ID NO: 71, a conservative modified variant of SEQ ID NO: 71 or a variant of SEQ ID NO: 71 comprising a standard structure class of 1 comprising the amino acid sequence of the variant of SEQ ID NO: 80; CDRH1 comprises SEQ ID NO: 80, the variant of SEQ ID NO: 80, a conservative modified variant of SEQ ID NO: 80 or a variant of SEQ ID NO: 80 comprising a standard structure class of 1 comprising the amino acid sequence of the variant of SEQ ID NO: 89; CDRH2 comprises SEQ ID NO: 89, the variant of SEQ ID NO: 89, SEQ ID NO: 8 9 a conservative modified variant of SEQ ID NO: 89 and the amino acid sequence of a variant of SEQ ID NO: 89 comprising a standard structure class of 2B comprising the amino acid sequence of the variant of SEQ ID NO: 98; CDRH3 comprises SEQ ID NO: 98, the variant of SEQ ID NO: 98, the conservative modified variant of SEQ ID NO: 98 or the amino acid sequence of a variant of SEQ ID NO: 98 comprising a standard structure class of 4 comprising.

[0037] In still other embodiments of mammalian antibodies (including canineized antibodies), CDRL1 is SEQ ID NO: 54, a variant of SEQ ID NO: 54, a conservative modified variant of SEQ ID NO: 54 or a variant of SEQ ID NO: 54 comprising a standard structure class of 1 comprising the amino acid sequence of a variant of SEQ ID NO: 54; CDRL2 is SEQ ID NO: 63, SEQ ID NO: 63 a variant of SEQ ID NO: 63, a conservative modified variant of SEQ ID NO: 63 or a variant of SEQ ID NO: 63 comprising a standard structure class of 1 comprising the amino acid sequence of a variant of SEQ ID NO: 63; CDRL3 is SEQ ID NO: 72, SEQ ID NO: 72 a variant of SEQ ID NO: 72, a conservative modified variant of SEQ ID NO: 72 or a variant of SEQ ID NO: 72 comprising a standard structure class of 1 comprising the amino acid sequence of a variant of SEQ ID NO: 81; CDRH1 is SEQ ID NO: 81, SEQ ID NO: 81 a variant of SEQ ID NO: 81, a conservative modified variant of SEQ ID NO: 81 or a variant of SEQ ID NO: 81 comprising a standard structure class of 1 comprising the amino acid sequence of a variant of SEQ ID NO: 90; CDRH2 is SEQ ID NO: 90, SEQ ID NO: 90 a conservative modified variant of SEQ ID NO: 90 and the amino acid sequence of a variant of SEQ ID NO: 90 comprising a standard structure class of 1 Includes sequence; CDRH3 is sequence number 99, variant of sequence number 99, conservative sequence number 99 Includes the amino acid sequence of the modified mutant or the mutant of SEQ ID NO: 99, which includes 13 standard structural classes. Hmm. In this particular embodiment of this type, the antibody (or its antigen-binding fragment) (T) is canine interleukin 4 receptor α (IL-4R α When the antibody binds to the sequence number In the amino acid sequence of sequence number 157 or sequence number 158, or in sequence number 157 and sequence number 158 At least one amino acid residue, preferably 2 to 5 amino acids, in both 158 and 158. It binds to a residue, and / or more preferably 3 to 8 or more amino acid residues.

[0038] In yet another embodiment of mammalian antibodies (including canine antibodies), CDRL1 is sequence number Variant of sequence number 55, variant of sequence number 55, conservative modified variant of sequence number 55, or standard structure of 2A Includes the amino acid sequence of the variant in SEQ ID NO: 55, which includes the class; CDRL2 is in SEQ ID NO: 64. Includes variants of SEQ ID NO: 64, conserved modified variants of SEQ ID NO: 64, or standard structural classes of 1. Includes the amino acid sequence of the variant of SEQ ID NO: 64; CDRL3 is SEQ ID NO: 73, SEQ ID NO: 7 SEQ ID NOs: 3 variants, the conserved modified variant of SEQ ID NO: 73, or the standard structure class of SEQ ID NOs: 1 Includes amino acid sequences of 73 variants; CDRH1 is a variant of SEQ ID NO: 82, SEQ ID NO: 82 , a conserved modified variant of SEQ ID NO: 82 or a variant of SEQ ID NO: 82 containing a standard structural class of 1 Includes the amino acid sequence of the body; CDRH2 is a variant of SEQ ID NO: 91, SEQ ID NO: 91, SEQ ID NO: The mutant of SEQ ID NO: 91, including 91 conserved modified mutants and the standard structure class of 2A. Contains a noacid sequence; CDRH3 is a variant of SEQ ID NO 100, SEQ ID NO 100, SEQ ID NO 10 Amino of the mutant of SEQ ID NO: 100, which includes 0 conservative modified mutants or 6 standard structural classes. Contains acid sequences.

[0039] In yet another embodiment of mammalian antibodies (including canine antibodies), CDRL1 is sequence number Syntax 129, a variant of SEQ ID NO: 129, a conservative modified variant of SEQ ID NO: 129, or a standard variant of 6. Includes the amino acid sequence of the variant of SEQ ID NO: 129, which includes the structural class; CDRL2 is SEQ ID NO: 132, a variant of sequence number 132, a conserved modified variant of sequence number 132, or a standard variant of 1 Includes the amino acid sequence of the variant of SEQ ID NO: 132, which contains the symposium class; CDRL3 is SEQ ID NO: 1 35, mutant of SEQ ID NO: 135, conservative modified mutant of SEQ ID NO: 135 or standard structure of 1 Includes the amino acid sequence of the variant in SEQ ID NO: 135, which includes the class; CDRH1 is SEQ ID NO: 14 0, a variant of SEQ ID NO: 140, a conserved modified variant of SEQ ID NO: 140, or a standard structure variant of 1. Includes the amino acid sequence of the variant of SEQ ID NO: 140 containing Ras; CDRH2 is SEQ ID NO: 144 , a mutant of SEQ ID NO: 144, a conserved modified mutant of SEQ ID NO: 144, and the standard structure of 3A Includes amino acid sequence of variant of SEQ ID NO: 144, which contains Ras; CDRH3 is SEQ ID NO: 149 , a mutant of SEQ ID NO: 149, a conserved modified mutant of SEQ ID NO: 149, or 15 standard structural variants This includes the amino acid sequence of a variant of SEQ ID NO: 149 containing Ras. This particular embodiment of this type. In this case, the antibody (or its antigen-binding fragment) is received by canine interleukin 4 receptor Condition α(IL-4R α When binding to ), the antibody is equivalent to SEQ ID NO: 127 or SEQ ID NO: 128 In the amino acid sequence, or in both SEQ ID NO: 127 and SEQ ID NO: 128, at least One more amino acid residue, preferably 2 to 5 amino acid residues, and / or more preferably It binds to 3 to 8 or more amino acid residues.

[0040] In yet another embodiment of mammalian antibodies (including canine antibodies), CDRL1 is sequence number Variant 130, variant of SEQ ID NO. 130, conservative modified variant of SEQ ID NO. 130, or standard variant 6. Includes the amino acid sequence of the variant of SEQ ID NO: 130, which includes the structural class; CDRL2 is SEQ ID NO: 133, a variant of sequence number 133, a conserved modified variant of sequence number 133, or a standard variant of 1 Includes the amino acid sequence of the variant of SEQ ID NO: 133, which contains the symposium class; CDRL3 is SEQ ID NO: 1 36, mutant of SEQ ID NO: 136, conservative modified mutant of SEQ ID NO: 136 or standard structure of 1 Includes the amino acid sequence of the variant in SEQ ID NO: 136, which includes the class; CDRH1 is SEQ ID NO: 14 1. A variant of SEQ ID NO: 141, a conserved modified variant of SEQ ID NO: 141, or a standard structural variant of 1. Includes amino acid sequence of the variant of SEQ ID NO: 141 containing Ras; CDRH2 is SEQ ID NO: 145 , a mutant of SEQ ID NO: 145, a conservative modified mutant of SEQ ID NO: 145, and the standard structure of 2A Includes the amino acid sequence of the variant of SEQ ID NO: 145, which contains Ras; CDRH3 is SEQ ID NO: 150 , a variant of SEQ ID NO: 150, a conservative modified variant of SEQ ID NO: 150, or 10 standard structural variants This includes the amino acid sequence of the variant of SEQ ID NO: 150 containing Ras. This particular embodiment of this type. In this case, the antibody (or its antigen-binding fragment) is received by canine interleukin 4 receptor Condition α(IL-4R α When binding to ), the antibody is equivalent to SEQ ID NO: 158 or SEQ ID NO: 162 In the amino acid sequence, or in both SEQ ID NO: 158 and SEQ ID NO: 162, One more amino acid residue, preferably 2 to 5 amino acid residues, and / or more preferably It binds to 3 to 8 or more amino acid residues.

[0041] In yet another embodiment of mammalian antibodies (including canine antibodies), CDRL1 is sequence number Syntax 129, a variant of SEQ ID NO: 129, a conservative modified variant of SEQ ID NO: 129, or a standard variant of 6. Includes the amino acid sequence of the variant of SEQ ID NO: 129, which includes the structural class; CDRL2 is SEQ ID NO: 134, a variant of sequence number 134, a conserved modified variant of sequence number 134, or a standard variant of 1 Includes the amino acid sequence of the variant of SEQ ID NO: 134, which contains the symposium class; CDRL3 is SEQ ID NO: 1 37, mutant of SEQ ID NO: 137, conservative modified mutant of SEQ ID NO: 137 or standard structure of 1 Includes the amino acid sequence of the variant in SEQ ID NO: 137, which includes the class; CDRH1 is SEQ ID NO: 14 0, a variant of SEQ ID NO: 140, a conserved modified variant of SEQ ID NO: 140, or a standard structure variant of 1. Includes the amino acid sequence of the variant of SEQ ID NO: 140, which contains Ras; CDRH2 is SEQ ID NO: 146 , a mutant of SEQ ID NO: 146, a conserved modified mutant of SEQ ID NO: 146, and the standard structure of 3A Includes the amino acid sequence of the variant of SEQ ID NO: 146, which contains Ras; CDRH3 is SEQ ID NO: 151 , a variant of sequence number 151, a conserved modified variant of sequence number 151, or 15 standard structural variants This includes the amino acid sequence of a variant of SEQ ID NO: 151 containing Ras. This particular embodiment of this type. In this case, the antibody (or its antigen-binding fragment) is received by canine interleukin 4 receptor Condition α(IL-4R α When binding to ), the antibody is equivalent to SEQ ID NO: 125 or SEQ ID NO: 126 In the amino acid sequence, or in both SEQ ID NO: 125 and SEQ ID NO: 126, at least One more amino acid residue, preferably 2 to 5 amino acid residues, and / or more preferably It binds to 3 to 8 or more amino acid residues.

[0042] In yet another embodiment of mammalian antibodies (including canine antibodies), CDRL1 is sequence number Variant 131, variant of sequence number 131, conservative modified variant of sequence number 131, or standard variant 3. Includes the amino acid sequence of the variant of SEQ ID NO: 131, which includes the structural class; CDRL2 is SEQ ID NO: 60, a variant of SEQ ID NO: 60, a conservative modified variant of SEQ ID NO: 60, or a standard structural variant of 1. Includes the amino acid sequence of the variant of SEQ ID NO: 60, which contains; CDRL3 is SEQ ID NO: 138, The mutant in sequence number 138, the conserved modified mutant in sequence number 1385, or the three standard structural classes. Includes the amino acid sequence of the variant of SEQ ID NO: 138; CDRH1 is SEQ ID NO: 142, The mutant of sequence number 142, the conserved modified mutant of sequence number 142, or the standard structure class of 1 Includes the amino acid sequence of the variant in SEQ ID NO: 142; CDRH2 is SEQ ID NO: 147, sequence The mutant of number 147, the conserved modified mutant of sequence number 147, and the standard structural class of 3A Includes the amino acid sequence of the variant in SEQ ID NO: 147; CDRH3 is SEQ ID NO: 152, sequence Includes mutant number 152, conserved modified mutant of sequence number 152, or standard structural class 9. This includes the amino acid sequence of the variant of SEQ ID NO: 152. In this particular embodiment of this type, The antibody (or its antigen-binding fragment) is found to be a canine interleukin 4 receptor α IL-4R α When the antibody binds to the sequence, the antibody is sequence number 154 or sequence number 155 or sequence At least one amino acid residue in number 156 or any combination thereof, preferred 2 to 5 amino acid residues, and / or more preferably 3 to 8 or more amino acid residues To bond to a base.

[0043] In yet another embodiment of mammalian antibodies (including canine antibodies), CDRL1 is sequence number Syntax 129, a variant of SEQ ID NO: 129, a conservative modified variant of SEQ ID NO: 129, or a standard variant of 6. Includes the amino acid sequence of the variant of SEQ ID NO: 129, which includes the structural class; CDRL2 is SEQ ID NO: 132, a variant of sequence number 132, a conserved modified variant of sequence number 132, or a standard variant of 1 Includes the amino acid sequence of the variant of SEQ ID NO: 132, which contains the symposium class; CDRL3 is SEQ ID NO: 1 39, mutant of SEQ ID NO: 139, conservative modified mutant of SEQ ID NO: 139 or standard structure of 1 Includes the amino acid sequence of the variant in SEQ ID NO: 139, which includes the class; CDRH1 is SEQ ID NO: 14 3. A mutant of SEQ ID NO: 143, a conserved modified mutant of SEQ ID NO: 143, or a standard structure variant of SEQ ID NO: 143. Includes the amino acid sequence of the variant of SEQ ID NO: 143 containing Ras; CDRH2 is SEQ ID NO: 148 , a mutant of SEQ ID NO: 148, a conservative modified mutant of SEQ ID NO: 148, and the standard structure of 3A Includes the amino acid sequence of the variant of SEQ ID NO: 148 containing Ras; CDRH3 is SEQ ID NO: 153 , a mutant of SEQ ID NO: 153, a conserved modified mutant of SEQ ID NO: 153, or 13 standard structural variants This includes the amino acid sequence of a variant of SEQ ID NO: 153 containing Ras. This particular embodiment of this type. In this case, the antibody (or its antigen-binding fragment) is received by canine interleukin 4 receptor Condition α(IL-4R α When binding to the antibody, the antibody is equivalent to SEQ ID NO: 159 or SEQ ID NO: 160. or at least one amino acid residue in SEQ ID NO: 161 or any combination thereof Preferably 2 to 5 amino acid residues, and / or more preferably 3 to 8 or more amino acid residues. It binds to amino acid residues.

[0044] This invention relates to the canine interleukin-4 receptor alpha (IL-4Rα ) specifically binds to This comprises an antibody and its antigen-binding fragment. In a particular embodiment of this type, The antibody and its antigen-binding fragment are canine IL-4R α Combined with InuIL- Dog IL-4R to IL-13 and / or IL-13 α It blocks the bond. As mentioned above, The detached mammalian antibody or its antigen-binding fragment is a canine antibody or its canine antibody It may be a primordial binding fragment. In other embodiments, isolated mammalian antibodies or The antigen-binding fragment is a mouse antibody or its mouse antigen-binding fragment. It's possible.

[0045] The canine antibody or its canine antigen-binding fragment of the present invention may include a hinge region. In this particular embodiment, the hinge region is the amino acid sequence of SEQ ID NO: 101. This includes. In another embodiment, the hinge region has the amino acid sequence of SEQ ID NO: 102 In another embodiment, the hinge region is the amino acid sequence of SEQ ID NO: 103. This includes. In yet another embodiment, the hinge region is the amino acid distribution of SEQ ID NO: 104 Includes columns.

[0046] In one embodiment, the canine antibody or its antigen-binding fragment is sequence number It contains a heavy chain containing the amino acid sequence of no. 164. In a particular embodiment of this type, The heavy chain is encoded by the nucleotide sequence of SEQ ID NO: 163. In other embodiments, The canine antibody or its antigen-binding fragment is the amino acid sequence of SEQ ID NO: 166. Includes a heavy chain containing . In a particular embodiment of this type, the heavy chain is SEQ ID NO: 165 Encoded by the nucleotide sequence. In yet another embodiment, the canine antibody The antigen-binding fragment contains a heavy chain that includes the amino acid sequence of SEQ ID NO: 168. In this particular embodiment, the heavy chain is the nucleotide sequence of SEQ ID NO: 167 This is coded by. In certain embodiments of such types, the canine antibody (ma (or its antigen-binding fragment) is the canine interleukin-4 receptor α (IL-4R α ) When binding, the antibody is also SEQ ID NO: 154 or SEQ ID NO: 155 or SEQ ID NO: 156 This is at least one amino acid residue in any combination thereof, preferably 2 to 5 It binds to an amino acid residue, and / or more preferably to 3 to 8 or more amino acid residues.

[0047] In related embodiments, the canine antibody or its antigen-binding fragment is sequence number It contains a light chain having the amino acid sequence of no. 170. In a particular embodiment of this type, The light chain is encoded by the nucleotide sequence of SEQ ID NO: 169. In other embodiments, The canine antibody or its antigen-binding fragment is the amino acid sequence of SEQ ID NO: 172. Includes a light chain containing . In a particular embodiment of this type, the light chain is SEQ ID NO: 171 Encoded by the nucleotide sequence. In yet another embodiment, the canine antibody The antigen-binding fragment contains a light chain with the amino acid sequence of SEQ ID NO: 174. In this particular embodiment, the light chain is the nucleotide sequence of SEQ ID NO: 173. This is coded by. In certain embodiments of such types, the canine antibody (ma (or its antigen-binding fragment) is the canine interleukin-4 receptor α (IL-4R α ) When binding, the antibody is also SEQ ID NO: 154 or SEQ ID NO: 155 or SEQ ID NO: 156 This is at least one amino acid residue in any combination thereof, preferably 2 to 5 It binds to an amino acid residue, and / or more preferably to 3 to 8 or more amino acid residues.

[0048] The present invention further provides antibodies comprising such combinations of heavy and light chains. Specific implementations In this state, the heavy chain contains the amino acid sequence of SEQ ID NO: 164, and the light chain contains the amino acid sequence of SEQ ID NO: 170 The amino acid sequence includes the following: In a more detailed embodiment of this type, the heavy chain is the sequence number. Encoded by a sequence of 163 nucleotides, the light chain is the nucleotide sequence of SEQ ID NO: 169 Coded by a column. In other embodiments, the heavy chain is the amino acid sequence of SEQ ID NO: 166 The light chain includes a column containing the amino acid sequence of SEQ ID NO: 172. A more detailed implementation of this type. In terms of form, the heavy chain is encoded by the nucleotide sequence of SEQ ID NO: 165, and the light chain This is encoded by the nucleotide sequence of sequence number 171. In yet another embodiment, The heavy chain contains the amino acid sequence of SEQ ID NO: 168, and the light chain contains the amino acid sequence of SEQ ID NO: 174. Includes a column. In a more detailed embodiment of this type, the heavy chain is the nucleus of Sequence ID No. 167. Encoded by a rheotide sequence, the light chain is coded by the nucleotide sequence of SEQ ID NO: 173. They'll be killed.

[0049] In related embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 164, and the light chain comprises It contains the amino acid sequence of sequence number 172. In other embodiments, the heavy chain is sequence number 164. The light chain contains the amino acid sequence of SEQ ID NO: 174. In this state, the heavy chain contains the amino acid sequence of SEQ ID NO: 166, and the light chain contains the amino acid sequence of SEQ ID NO: 170 It contains the amino acid sequence of SEQ ID NO: 166. In yet another embodiment, the heavy chain is the amino acid sequence of SEQ ID NO: 166. It contains an acid sequence, and the light chain contains the amino acid sequence of SEQ ID NO: 174. In other embodiments, Therefore, the heavy chain contains the amino acid sequence of SEQ ID NO: 168, and the light chain contains the amino acid sequence of SEQ ID NO: 170. It includes an acid sequence. In other embodiments, the heavy chain includes the amino acid sequence of SEQ ID NO: 168. The light chain contains the amino acid sequence of SEQ ID NO: 172.

[0050] In certain embodiments of that type, the canine antibody (or its antigen-binding properties) Fragment) is canine interleukin 4 receptor α (IL-4R α When bonding with the The body is sequence number 154, sequence number 155, sequence number 156, or any combination thereof. At least one amino acid residue in the combination, preferably 2 to 5 amino acid residues, and It binds to / or more preferably 3 to 8 or more amino acid residues.

[0051] Therefore, the present invention further relates to the canine interleukin 4 receptor α (IL-4R α ) unique An isolated mammalian antibody or its antigen-binding fragment (canine antibody) that binds to it. The antibody provides (including its antigen-binding fragment), where the antibody is canine IL-4R α When joining, sequence number 125 or sequence number 126 or sequence number 127 or sequence Number 128 or Sequence ID 154 or Sequence ID 155 or Sequence ID 156 or Sequence ID Number 157 or Sequence ID 158 or Sequence ID 159 or Sequence ID 160 or Array At least one of the numbers 161 or sequence number 162 or any combination thereof Amino acid residues, preferably 2 to 5 amino acid residues, and / or more preferably 3 to It binds to eight or more amino acid residues. In certain embodiments, the antibody or its antigen The binding fragment is canine IL-4R α Binds to canine interleukin 4 and canine IL -4R α It blocks the bond.

[0052] The present invention further includes 1 × 10 -12 Dissociation constants (Kd) lower than M (e.g., 1 × 10⁻⁶) -13 Dog IL-4R (M or below) α Mammalian antibodies or their antigen-binding fragments that bind to Provided. In certain embodiments, the mammalian antibody or its antigen-binding fragment. is 1 x 10 -5 M~1×10 -12 The dissociation constant of M is for canine IL-4R α Combine with. More details In a detailed embodiment, the mammalian antibody or its antigen-binding fragment is 1 × 10 - 7 M~1×10 -11 The dissociation constant of M is for canine IL-4R α Combine. Further detailed implementation In terms of morphology, the mammalian antibody or its antigen-binding fragment is 1 × 10 -8 M~1 ×10 -11 The dissociation constant of M is for canine IL-4R α It is combined with a more detailed embodiment. In this case, the mammalian antibody or its antigen-binding fragment is 1 × 10 -8 M~1×10 - 10 The dissociation constant of M is for canine IL-4R α Combine.

[0053] This invention also includes 1 × 10 7 M-1 s -1 with a larger on-rate (k on ) binds to canine IL-4R α to provide a mammalian antibody or an antigen-binding fragment thereof. In certain embodiments, the mammalian antibody or an antigen-binding fragment thereof has an on-rate (k 2 M -1 s -1 ~1×10 7 M -1 s -1 and binds to canine IL-4R with an on-rate (k on ) of α . In more specific embodiments, the mammalian antibody or an antigen-binding fragment thereof has an on-rate of 1×10 1×10 3 M -1 s -1 ~1×10 6 M -1 s -1 and binds to canine IL-4R. In even more specific embodiments, the mammalian antibody or an antigen-binding fragment α thereof has an on-rate of 1×10 1×10 3 M -1 s -1 ~1×10 5 M -1 s -1 and binds to canine IL-4R. In even more specific embodiments, the mammalian antibody or an antigen-binding fragment α thereof has an on-rate of 1×10 4 M -1 s -1 ~1×10 5 M -1 s -1 and binds to canine IL- 4R α .

[0054] The present invention further provides an off-rate (k slower than 1×10 -7 s -1 ).off ) Dog I L-4R α Provides a mammalian antibody or its antigen-binding fragment that binds to a specific target. In this embodiment, the mammalian antibody or its antigen-binding fragment is 1 × 10 -3 s -1 ~1 × 10 -8 s -1 At the off speed, the IL-4R α It connects to. More detailed implementation In this state, the mammalian antibody or its antigen-binding fragment is 1 × 10 -4 s -1 ~ 1 x 10 -7 s -1 At the off speed, the IL-4R α Combined. Further detailed embodiments In this case, the mammalian antibody or its antigen-binding fragment is 1 × 10 -5 s -1 ~1 ×10 -7 s -1 At the off speed, the IL-4R α Combine.

[0055] In certain embodiments, the mammalian antibody (including a chimeric antibody) of the present invention is IL-4R α It blocks the binding of canine IL-4 to it. In a more detailed embodiment, the antibody is 1 × 10 -8 M~1×10 -9 IL-4R at M or even lower concentrations with a minimum EC50 α Dogs It blocks the binding of IL-4. In a more detailed embodiment, EC50 is 5 × 10 - 9 M~5×10 -13 M. In a more detailed embodiment, EC50 is 5 × 1 0 -9 M~5×10 -11 It is M.

[0056] In related embodiments, the mammalian antibody or its antigen-binding fragment is type I By IL-4 and / or IL-13 that bind to the type II IL-4 receptor Negatively attenuates (e.g., suppresses) the cellular signaling pathway mediated by it. Specific implementation Morphologically, the mammalian antibody or its antigen-binding fragment is used in animals to scratch It improves pruritic inflammatory skin diseases, such as atopic dermatitis. In a more detailed embodiment, it improves these conditions. In this embodiment, the animal subject is a dog. In the related embodiment, the animal subject is a cat.

[0057] Therefore, any antibody of the present invention possesses these properties, namely, canine IL-4R α Regarding The aforementioned dissociation constant, canine IL-4R α The aforementioned on-rate of binding to the antibody-dog IL4R α The aforementioned off-rates for dissociation from the binding complex, type I and / or type II I Cellular signaling mediated by IL-4 and / or IL-13 binding to the L-4 receptor Suppression of signaling pathways, or pruritic inflammatory skin diseases in animals, such as atopic dermatitis. It may show one, two, three, four, or all of the improvements in dermatitis.

[0058] As described above, the antibody of the present invention, comprising the antibody (and its antigen-binding fragment) (and its antigen-binding fragment) is a monoclonal antibody (and its antigen-binding fragment) (glutination), mammalian antibodies (and their antigen-binding fragments), for example, mice (mouse) ) Antibodies (and their antigen-binding fragments), canine antibodies (and their antigen-binding fragments) (This includes a canine mouse antibody (and its antigen-binding fragment)) This is possible, and in one embodiment, the antibody (and its antigen-binding fragment) ) is isolated.

[0059] The present invention further relates to a nucleic acid (isolated) that encodes any one of the light chains of the canine antibody of the present invention. The present invention provides a nucleic acid (containing the above). Similarly, the present invention provides any one of the heavy chains of the canine antibody of the present invention. It provides isolated nucleic acids that encode two things.

[0060] The present invention further relates to an expression vector comprising one or more nucleic acids (including isolated nucleic acids) of the present invention. The present invention further provides a host cell containing one or more of the expression vectors of the present invention.

[0061] In certain embodiments, the antibody is a recombinant antibody or its antigen-binding fragment. Yes. In related embodiments, the variable heavy chain domain and the variable light chain domain are single-chain anti They are connected by flexible linkers to form a body.

[0062] In certain embodiments, the antibody or antigen-binding fragment is a Fab fragment. In other embodiments, the antibody or antigen-binding fragment is Fab's. It is a fragment. In other embodiments, the antibody or antigen-binding fragment is ( This is the Fab')2 fragment. In yet another embodiment, the antibody or antigen binds to it. The sex fragment is a diabody. In certain embodiments, the antibody or antigen binds to it. The sex fragment is a domain antibody. In certain embodiments, the antibody or antigen is The compatible fragment is a single-domain antibody.

[0063] In certain embodiments, canine-like mice are treated with anti-canine IL-4R α Antibody or antigen-binding Ragment is a type I and / or II disorder in the animal being treated (e.g., dogs). Cellular signatures mediated by IL-4 and / or IL-13 binding to type IL-4 receptors The Nal transmission pathway is negatively attenuated. In a more detailed embodiment, the canine mouse of the present invention Anti-canine IL-4R α Administration of antibodies or antigen-binding fragments is used in animals being treated. For example, it works to improve one or more symptoms of atopic dermatitis in dogs.

[0064] This invention further relates to canine mice and anti-canine IL-4R α Isolation of antibodies or parts thereof that encode an antibody Provides nucleic acids that have been processed. In related embodiments, such antibody or antigen-binding fungible The compound is used in the manufacture of pharmaceuticals for the treatment of atopic dermatitis in dogs. It may be used. Alternatively or in combination, the present invention may be used for diagnostic purposes, with the antibodies or The use of any of the antibody fragments is provided. Further embodiments provide further details. A kit containing either a canine antibody or an antigen-binding fragment disclosed in the details. provide.

[0065] In a further embodiment, the canine mouse anti-canine IL-4R of the present invention α Antibodies This expression vector contains an isolated nucleic acid encoding one of the antigen-binding fragments. The present invention also provides a host comprising any of the expression vectors described herein. Regarding cells. In certain embodiments, these nucleic acids, expression vectors or Polypeptides are useful in antibody production methods. The present invention further relates to SEQ ID NO: 125 and This is sequence number 126 or sequence number 127 or sequence number 128 or sequence number 154 This is sequence number 155 or sequence number 156 or sequence number 157 or sequence number 158 This is the sequence number 159, or sequence number 160, or sequence number 161, or sequence number 162. A peptide consisting of 80 or fewer amino acid residues containing a amino acid sequence (isolated antigen peptide) Provides (including) the peptide (isolated antigen peptide). In related embodiments, the peptide (isolated antigen peptide) (including) SEQ ID NO: 125 or SEQ ID NO: 126 or SEQ ID NO: 127 or SEQ ID NO: 1 28 or SEQ ID NO: 154 or SEQ ID NO: 155 or SEQ ID NO: 156 or SEQ ID NO: 1 57 or SEQ ID NO: 158 or SEQ ID NO: 159 or SEQ ID NO: 160 or SEQ ID NO: 1 It consists of 60 or fewer amino acid residues containing the amino acid sequence of 61 or SEQ ID NO: 162. In the linked embodiment, the peptide (including the isolated antigen peptide) is SEQ ID NO: 12 5 or SEQ ID NO: 126 or SEQ ID NO: 127 or SEQ ID NO: 128 or SEQ ID NO: 15 4 or SEQ ID NO: 155 or SEQ ID NO: 156 or SEQ ID NO: 157 or SEQ ID NO: 15 8 or SEQ ID NO: 159 or SEQ ID NO: 160 or SEQ ID NO: 161 or SEQ ID NO: 16 It consists of 10 to 45 amino acid residues containing the amino acid sequence 2. In other embodiments, Therefore, the peptide (including the isolated antigen peptide) is SEQ ID NO: 125 or SEQ ID NO: 126 or SEQ ID NO: 127 or SEQ ID NO: 128 or SEQ ID NO: 154 or SEQ ID NO: 155 or SEQ ID NO: 156 or SEQ ID NO: 157 or SEQ ID NO: 158 or SEQ ID NO: Amino acid sequence of 159 or SEQ ID NO: 160 or SEQ ID NO: 161 or SEQ ID NO: 162 It consists of 5 to 25 amino acid residues, either from or including them.

[0066] The present invention further includes Sequence ID 125 or Sequence ID 126 or Sequence ID 127 or Sequence ID 127 or Sequence ID Number 128 or Sequence ID 154 or Sequence ID 155 or Sequence ID 156 or Sequence ID Number 157 or Sequence ID 158 or Sequence ID 159 or Sequence ID 160 or Array 80%, 85%, 90%, 95% for the amino acid sequence of number 161 or sequence number 162 They are % or 100% identical, and the isolated mammalian antibody or its antigen-binding fraction of the present invention is identical. Antigen peptide consisting of 80 or fewer amino acid residues, including the amino acid sequence that binds to the pigment. (including isolated peptides) is provided. In related embodiments, the antigen peptide ( (Containing isolated antigen peptides) SEQ ID NO: 125 or SEQ ID NO: 126 or sequence number Number 127 or Sequence ID 128 or Sequence ID 154 or Sequence ID 155 or Sequence Number Sequence No. 156 or Sequence No. 157 or Sequence No. 158 or Sequence No. 159 or Sequence No. 80% of the amino acid sequence of number 160 or SEQ ID NO: 161 or SEQ ID NO: 162, 8 Isolated mammalian antibodies or their antigens that are 5%, 90%, 95%, or 100% identical. It consists of 60 or fewer amino acid residues, including an amino acid sequence that binds to a binding fragment. In other embodiments, the peptide is SEQ ID NO: 125 or SEQ ID NO: 126 or Column number 127 or sequence number 128 or sequence number 154 or sequence number 155 or Column number 156 or sequence number 157 or sequence number 158 or sequence number 159 or 80% for the amino acid sequence of sequence number 160 or sequence number 161 or 162 , 85%, 90%, 95%, or 100% identical, isolated mammalian antibodies or 5 to 25 amino acid sequences from or containing those sequences that bind to antigen-binding fragments. It consists of the amino acid residues. In certain embodiments, the mammalian antibody has a CDR of 4D8. Includes. In other embodiments, the mammalian antibody includes 11H2 CDR. Further embodiments In terms of form, the mammalian antibody contains a 4H3 CDR. In yet another embodiment, The mammalian antibody contains 11B6 CDR. In yet another embodiment, the mammalian antibody is In another embodiment, the mammalian antibody contains a 2E2 CDR. Includes.

[0067] The present invention further provides a fusion protein comprising any of the aforementioned peptides. Specific implementation Morphologically, the fusion protein is composed of such antigenic peptides and non-canine mammalian IgG antibodies. It includes the Fc region of the fusion protein. In a more detailed embodiment, the fusion protein is a non-canine mammal. It contains the Fc region of the IgG antibody. In one embodiment, the non-canine mammalian IgG antibody is It is weak IgG. In an alternative embodiment, the non-canine mammalian IgG antibody is human IgG. In other embodiments, the non-canine mammalian IgG antibody is equine IgG. In other embodiments, the non-canine mammalian IgG antibody is porcine IgG. Morphologically, the non-canine mammalian IgG antibody is bovine IgG.

[0068] In certain embodiments, the non-canine mammalian IgG antibody is IgG1. Other embodiments In this embodiment, the non-canine mammalian IgG antibody is IgG2a. Furthermore, in other embodiments... In this embodiment, the non-canine mammalian IgG antibody is IgG3. In yet another embodiment, the non The canine mammalian IgG antibody is IgG4. In other embodiments, the fusion protein is The present embodiment comprises one of the aforementioned antigen peptides and a maltose-binding protein. In this case, the fusion protein is a beta-galactosider of one of the antigen peptides. It contains a ze. In yet another embodiment, the fusion protein is one of the antigen peptides. It comprises either and glutathione S-transferase. In yet another embodiment, the The fusion protein comprises one of the aforementioned antigen peptides and thioredoxin. Further embodiments Morphologically, the fusion protein contains either of the antigen peptides and Gro EL. In another embodiment, the fusion protein is one of the antigen peptides and N Includes USA.

[0069] The present invention further comprises nucleic acids (single) encoding the antigen peptide and corresponding fusion protein of the present invention. The present invention also provides an expression vector containing these nucleic acids. The present invention provides a host cell containing one or more expression vectors of the present invention.

[0070] Furthermore, the present invention relates to the anti-canine IL-4R α Antibodies or their antigen-binding fragments , Inu IL-4R α Antigen peptides from (including isolated antigen peptides), the present invention NuIL-4R α A fusion protein containing antigen peptides from the present invention, the antigen fragment and / or or nucleic acids (including isolated nucleic acids) that encode fusion proteins, including such nucleic acids An expression vector or any combination thereof, and a pharmaceutically acceptable carrier or diluent. Includes a pharmaceutical composition containing the following:

[0071] Furthermore, the present invention is a method for negatively attenuating the activity of IL-4 and / or IL-13. This includes administering a therapeutically effective amount of such pharmaceutical composition to an animal requiring it. The method provides a method for treating atopic dermatitis in dogs. In one embodiment, the method provides a method for treating atopic dermatitis in dogs. It is used.

[0072] These and other aspects of the present invention are described in the "Brief Description of the Drawings" and "Detailed Description" below. This will be better understood by referring to [the relevant source].

[0073] Detailed explanation Various approaches to treating human Alzheimer's disease (AD) are currently being investigated in numerous clinical trials. [Malajian et al., New pathogenic and therapeutic ic paradigms in atopic dermatitis Cytoki This is outlined in ne, (2014). Some of these approaches are in Th2 The aim is to inhibit one or more signaling molecules / events that trigger cell development and activation. One direction of research in this area is key interleukin-driven materials of the Th2 pathway. This includes approaches to block the action of [unclear]. AD is a disease primarily controlled by Th2 [unclear]. Observations, as well as the key driving forces behind the development of Th2 cells, both IL-4 and IL-13 Based on accumulated data supporting the central role in the combined effects of the two, and IL-4 receptor This shows that the alpha chain is an essential receptor for signaling from both cytokines. Based on the data, the present invention relates to canine IL-4 and I to type I and type II IL-4 receptors. The binding of nu IL-13 is blocked, and then signals from both nu IL-4 and IL-13 are blocked. This paper describes the construction and characterization of monoclonal antibodies that inhibit signal transduction. These antibodies are As disclosed herein, atopic dermatitis and other diseases in companion animals It has therapeutic utility.

[0074] Abbreviation The following abbreviations are used throughout the detailed description and examples of the present invention.

[0075] ADCC antibody-dependent cytotoxicity; CDC complement-dependent cell injury; The immunoglobulin variable regions defined using the CDR Kabat numbering system. Complementarity determination areas within a region; CHO Chinese hamster ovaries; EC50: A concentration that yields 50% potency or binding; ELISA (Enzyme-Linked Immunosolvent Assay); FR antibody framework region: immunoglobulin variable region excluding the CDR region; HRP (Horseradish Peroxidase); IFN (Interferon); Concentration that results in a 50% inhibition of IC50; IgG (Immunoglobulin G) Kabat Elvin A. Kabat[Sequences of Protei ns of Immunological Interest, 5th Ed.Pub lic Health Service, National Institutes o Immunoglobulin developed by f Health, Bethesda, Md. (1991)] Brin alignment and numbering systems; mAb monoclonal antibody (also known as Mab or MAb); PCR (polymerase chain reaction); MES 2-(N-morpholino)ethanesulfonic acid; Mechanism of action of MOA; NHS normal human serum; PCR (polymerase chain reaction); PK (pharmacokinetics); TT tetanus toxoid; The V region is a segment of the IgG chain whose sequence is variable between different antibodies. It is the K region within the light chain. It extends to abat residue 109 and 113 within the heavy chain; VH immunoglobulin heavy chain variable region; VL (Variable Region of Immunoglobulin Light Chain); VK: Variable region of the immunoglobulin kappa light chain.

[0076] definition To facilitate understanding of this invention, several scientific and technical terms are specifically defined below. To do so. Unless otherwise defined elsewhere in this specification, all other terms used herein. The scientific and technical terms used here have meanings that are generally understood by those skilled in the art to which this invention pertains.

[0077] In this specification, including the attached claims, singular words are used in a manner that is clearly inconsistent with the context. Unless a shield is used, the corresponding target includes multiple objects.

[0078] "Activation" is used when it applies to cells or receptors, depending on the context or explicitly specified. Unless otherwise indicated, activation or treatment (treatment) of cells or receptors with ligands is not intended. It can taste. "Ligand" refers to natural and synthetic ligands, such as cytokines. This includes variants, analogs, mutant proteins, and antibody-derived binding compounds. "Land" also includes small molecules, such as cytokine peptide mimes and antibody peptides. Includes mimics. "Activation" is regulated by internal mechanisms and by external or environmental factors. This could mean the activation of the affected cells.

[0079] The "activity" of a molecule refers to its binding to a ligand or receptor, or its catalytic activity; The ability to stimulate gene expression or cell signaling, differentiation, or maturation; antigenic activity, other functions This can indicate or imply modulation of the activity of the molecule. The "activity" of a molecule refers to intercellular activity. Interactions, such as activity when modulating or maintaining adhesion, or the structure of cells. It can also mean activity in the formation of, for example, the cell membrane or cytoskeleton. "Activity" is relative. Activity, for example, [catalytic activity] / [mg protein], or [immune activity] / [mg protein] "Activity" can also refer to the concentration within a biological compartment, etc. This could mean modulation of the components.

[0080] "Administration" and "treatment (processing)" refer to the use of animals, such as dogs, experimental subjects, cells, and tissues. When applied to organs or biological fluids, the subject may be an animal, such as a dog, or a cell or tissue. This refers to contact between an exogenous drug, therapeutic substance, diagnostic agent, or composition and an organ or biological fluid. The processing of cells involves contact between the cells and the reagent, and between the reagent and the fluid in contact with the cells. This includes contact with reagents, diagnostic agents, binding compounds or In vitro and ex vivo treatment of cells by other cells, for example. It also means that the word "subject (test subject)" refers to any living organism, preferably an animal, more preferably This includes mammals (e.g., dogs, cats, or humans), most preferably dogs.

[0081] In this specification, for example, in the amino acid sequence of an antibody, "A" at another amino acid residue. "Substitution of amino acid residues" is synonymous with "substituting an amino acid residue with another amino acid residue." Yes, a specific amino acid residue at a specific position in the amino acid sequence is different from a different amino acid residue. This indicates that it is substituted by. Such substitutions can be specifically designed, i.e., For example, recombinant DNA technology can be used to convert alanine to serine at a specific position within the amino acid sequence. It is possible to intentionally replace it. Alternatively, through a more natural selection process, for example, Antibodies produced by cells contain the antigen (e.g., an epitope or a part thereof). Based on its ability to bind to a given region on and / or the CDR that is to be replaced, The antibody contains a specific CDR that has the same standard structure, and the specific amino acid residues of the antibody also It is possible to substitute one or more amino acid residues in a chain of amino acid residues. Substitutions can result in "mutant" CDRs and / or mutant antibodies.

[0082] "To treat (process)" or "treatment (process)" refers to a therapeutic substance (for example, the present invention) A composition containing either an antibody or an antigen-binding fragment, wherein the substance has therapeutic activity Dogs or patients exhibiting one or more disease symptoms or suspected to have the disease are targeted for internal treatment. Alternatively, it means administering it externally.

[0083] Typically, the substance reduces such symptoms to any clinically measurable degree. Induction of contracture or suppression of the progression of such symptoms, resulting in one or more cases in the treatment population or group. It is administered in a dose effective to alleviate and / or improve the symptoms of any particular disease. The amount of a therapeutic substance that is effective in alleviating disease symptoms (also called the "therapeutic effective dose") is The patient's (e.g., dog) condition, age, and weight, as well as the ability to elicit a desired response in the subject. This can vary depending on factors such as the ability of the pharmaceutical composition. Disease symptoms may be reduced or improved. Whether or not a veterinarian or other skilled medical professional can assess the severity or progression of the symptoms. It can be evaluated by any of the clinical measurements typically used by the provider. The embodiment (e.g., treatment method or product) reduces the symptoms of the target disease in all subjects. It may not always be effective in this case, but it is known in the art. Statistical tests such as the Student t-test and chi-squared tests. 2 Tests, Mann and Hoy Whitney's U test, Kruskal-Wa llis) test (H test), Jonckheere-Terpstra Statistics determined by the pstra test and the Wilcoxon test. The target disease symptoms should be alleviated in a statistically significant number of subjects.

[0084] "Treatment (processing)" is when it is suitable for humans, veterinary subjects (e.g., dogs) or research subjects. When used, it means therapeutic treatment, research, and diagnostic application. "Treatment (treatment)" means, That is a human, a veterinary subject (e.g., a dog), or a research subject, or cells, tissues or When applied to organs, the target animals may be dogs or other animals, cells, tissues, physiological compartments or This includes contact of the antibody or antigen-binding fragment of the present invention with a physiological fluid.

[0085] In this specification, the term "dog" refers to all domesticated dogs, unless otherwise specified. Canis lupus familiaris This includes Canis familiaris.

[0086] In this specification, the term "cat" refers to any member of the Felidae family. It tastes good. Members of this family include wild, zoo, and domestic members, such as cats (Feli). Any member of the subfamily nae, e.g., cat, lion, tiger, puma, jaguar, rhinoceros Leopard, snow leopard, panther, North American mountain n) Lion, cheetah, lynx, bobcat, caracal or any of them This includes crossbreeds. Cats also include domestic cats, purebred and / or mixed companion cats, and show cats (show cats). This includes cats used for animal husbandry, laboratory cats, cloned cats, and wild or feral cats.

[0087] In this specification, the term "dog frame" is defined as a CDR residue. This refers to the amino acid sequences of the heavy and light chains of canine antibodies, excluding the hypervariable region residues. In many embodiments, with respect to canine antibodies, the amino acid sequence of natural canine CDR is... The chain is substituted with a corresponding foreign CDR (e.g., derived from a mouse antibody). For example, as illustrated below, the conformation of exogenous CDRs in canine antibodies To maintain the function and / or modify the Fc function, the heavy chain of the canine antibody The light chain may contain several foreign non-CDR residues.

[0088] Inu IL-4R α This is the sequence number 2 [sequence number 4 if it does not contain a signal sequence] It has been found to contain a mino acid sequence. In certain embodiments, canine IL-4R α teeth , containing the nucleotide sequence of SEQ ID NO: 1 [SEQ ID NO: 3 if it does not contain a signal sequence] Encoded by nucleic acids. Canine IL-4Rα For example, a non-conserved region contains a conserved mutation. They may differ in some respects, but the dog IL-4R α This is sequence number 2 [that does not contain a signal sequence] [SEQ ID NO: 4] contains the amino acid sequence of canine IL-4R α It has essentially the same biological function as .

[0089] Cytokines IL-4 and IL-13 are involved in various allergic diseases in humans and animals. It is associated with the pathogenesis of diseases (including asthma and atopic dermatitis). IL-4 receptor Since the alpha chain is an essential receptor for signal transduction from these cytokines, The invention is IL-4R α Blocking the binding of canine IL-4 and canine IL-13 to canine IL Production of monoclonal antibodies that suppress signal transduction from both IL-4 and IL-13. The characteristics are described. Therefore, these antibodies are disclosed herein. It has utility in the treatment of atopic dermatitis and other diseases in companion animals. Also, Inu IL-4R α The biological function of is, for example, specifically bound by the antibody of this disclosure. This may involve having an epitope within the extracellular domain.

[0090] Individual dogs IL-4R α The amino acid sequence generally includes the amino acid sequence of SEQ ID NO: 4. NuIL-4R α It is at least 90% identical to [another substance]. In some cases, it is canine IL-4R α This is canine IL-4R containing the amino acid sequence of SEQ ID NO: 4. α At least 95% or Furthermore, they are at least 96%, 97%, 98%, or 99% identical. In one embodiment, In the case of the dog IL-4Rα The amino acid sequence contains the amino acid sequence of SEQ ID NO: 4, canine IL- 4R α It shows a difference of 10 amino acids or less compared to [another example]. In one embodiment, canine IL- 4R α The amino acid sequence is the same as that of Canine IL-4R, which contains the amino acid sequence of SEQ ID NO: 4. α Compared to 5 Differences of less than one amino acid, or even less than four, three, two, or one amino acid, may be observed. Identity The percentage (%) can be determined as described below.

[0091] The term "immune response" refers to, for example, lymphocytes, antigen-presenting cells, phagocytic cells, granulocytes, and pre- Soluble macromolecules produced by cells or the liver (including antibodies, cytokines, and complement) The action of (m) against cancer cells, cells or tissues infected with pathogens, or invasive pathogens Selective damage, destruction, or removal of them from the body of a mammal (e.g., the body of a dog) It means the effect it brings about.

[0092] Anti-canine IL-4R α antibody This invention relates to Canine IL-4R α Isolated antibodies that bind to (especially mouse anti-canine IL-4R) α Antibodies and their canine antibodies) or their antigen-binding fragments, and such anti Provides the use of the body or fragments thereof. In certain embodiments, canine IL-4 R α Binds to, canine IL-4R α The ligand for this is canine IL-4 or IL-13 It has been shown to block the binding of one or more of the mouse anti-canine IL-4R α From antibodies Mouse anti-canine IL-4R α We provide CDRs. These CDRs are modified with canine antibodies. Inserted into the frame, canine-like mouse anti-canine IL-4R α It is possible to obtain antibodies.

[0093] The term "anti-canine IL-4R" as used herein α "Antibodies" are (for example, mice or rabbits) (In mammals such as) Canine IL-4R α Canine IL-4R produced against α In particular This refers to antibodies that bind heteromorphically. "Canine IL-4R α (And especially the IL-4R dogs α ) Antibodies that "specifically bind" or "canine IL-4R" α polypeptide containing the amino acid sequence The antibody that specifically binds to canine IL-4R compared to other antigens. α Shows preferential binding to Although it is an antibody, this specificity does not require absolute binding specificity. Anti-canine IL-4R α antibody The dog IL-4R α It is considered "specific" because the binding occurs in dog samples. Without inappropriately interfering with the activity of other molecules, for example, in a diagnostic setting, false positive or Without causing undesirable results such as side effects in the treatment setting, the sample contains NuIL-4R α If it determines the presence of canine IL-4R α The activity This is a case where sex can be altered. Anti-canine IL-4R α The degree of specificity required for an antibody is the antibody It may depend on the intended use, but in any case, its suitability for the intended purpose Determined by compatibility. Derived from the antibody of the assumed method, or from the antigen-binding site of the antibody. The binding compound is attached to either the antigen or its variant or mutant protein. At least twice as large an affinity for other antigens, preferably at least 10 Twice as large, more preferably at least 20 times larger, and most preferably at least 100 times larger They are joined with high affinity.

[0094] Given antigen sequence (in this case, canine IL-4R) α (The part of the amino acid sequence) The reason why antibodies specifically bind to lipeptides is because of canine IL-4R α amino It binds to polypeptides containing the relevant portion of the acid sequence, but not to canine IL-4R α that part of the said array This is the case when it does not bind to other canine proteins that lack the necessary component. For example, canine IL-4R α Includes The antibody that specifically binds to the polypeptide is canine IL-4R. α FLAG (registered trademark) tag It can bind to the attached form, but not to other FLAG® tagged canine proteins. No. The antibody, or a binding compound derived from the antigen-binding site of the antibody, will bind to the canine antigen or It can be said that it "specifically" binds to that variant or mutant protein because it , at least 10 times greater affinity than its affinity to any other tested canine antigen, More preferably at least 20 times larger, and even more preferably at least 100 times larger , having affinity for the canine antigen or its variant or mutant protein This is the case.

[0095] As used herein, the term "antibody" refers to any form of antibody that exhibits the desired biological activity. Therefore, it is used in the broadest sense, and in particular, monoculi that exhibit the desired biological activity. Ronal antibodies (including full-length monoclonal antibodies), polyclonal antibodies, multispecific antibodies Body (e.g., bispecific antibodies), canine antibodies, fully canine antibodies, chimeric antibodies and camel-derived antibodies. This includes, but is not limited to, single-domain antibodies. "Parent antibody" is intended to be... Modification of antibodies for use (e.g., canine modification of antibodies for use as canine therapeutic antibodies) These are antibodies obtained by the immune system's exposure to an antigen prior to the antigen's onset.

[0096] In this specification, "antibody fragment" or "antigen-binding fragment" refers to an antibody. The antigen-binding fragment, i.e., specifically binds to the antigen to which the full-length antibody binds. Antibody fragments that possess the ability to do so, for example, fragments that possess one or more CDR regions. It means "a point." Examples of antigen-binding fragments include Fab, Fab', and F(ab')2. and Fv fragments; diabodies; linear antibodies; single-chain antibody molecules, e.g., scFv; This also includes multispecific antibodies and nanobodies formed from antibody fragments. However, this is not limited to these.

[0097] "Fab Fragment" consists of one light chain and one heavy chain of carbon. H From 1 and the variable region It is constructed in such a way that the heavy chains of Fab molecules cannot form disulfide bonds with other heavy chain molecules. "Fab fragments" can be products of papain cleavage of antibodies.

[0098] The "crystallizable fragment" ("Fc") region is the C of the antibody. H 3 and C H 2 domains It contains two heavy chain fragments, which include 2 or more By disulfide bonds, and C H The three domains are bound together by hydrophobic interactions.

[0099] "Fab' fragment" is a single light chain, and V H Domain and C H 1 domain A portion or fragment of a heavy chain containing, and C H 1 and C H Area between two domains It contains, and as a result, interchain disulfides between the two heavy chains of the two Fab' fragments A bond can be formed to create an F(ab')2 molecule. The "F(ab')2 fragment" is , two light chains, and C H 1 and C H Two components containing the constant region between the two domains It contains heavy chains, and as a result, interchain disulfide bonds are formed between these two heavy chains. Therefore, the F(ab')2 fragment has a disulfide between its two heavy chains. It consists of two Fab' fragments linked by a bond. A "fragment" can be a product of pepsin cleavage of an antibody.

[0100] The "Fv region" includes variable regions from both the heavy and light chains, but lacks a steady region. .

[0101] The term "single-stranded Fv" or "scFv" antibody refers to the V in antibody. H and V L Domain Antibody fragments in which these domains are located within a single polypeptide chain. This means that, generally, Fv polypeptides can further form the desired structure for antigen binding using scFv. V H and V L Includes a polypeptide linker between domains [Pluckthun,THE PHARMACOLOGY OF MONOCLONA L ANTIBODIES, vol.113, edited by Rosenburg and Moore. Springer-Verlag, New York, pp.269-315(1994 );WO 88 / 01649;and US4,946,778 and US5, See 260,203.

[0102] In this specification, the term "canonical structure" refers to an antibody. Each of the hypervariable regions of the heavy and light chains is taken within the framework in which they exist. This refers to various local conformations. For each hypervariable region, there are a few standard structures (general There exist (represented by simple integers such as 1 or 2), and these are corresponding hyper Amino acid sequence of the variable region [in particular, the corresponding anti-canine IL-4R α Further information regarding the variable domain will be provided later. The amino acid sequence of that framework shown is in the environment (see Table 3 below). It can be predicted with high accuracy from (what you want to be done). These standard structures are the A Does modification of the amino acid sequence result in the retention or loss of its ability to bind to its antigen-binding partner? This could be decisive regarding [Chothia and Lesk, Canonic] al Structures for the hypervariable regi ons of immunoglobulins,J.Mol.Biol.196:90 1-917 (1987); Chothia et al., Conformation of imm unoglobulin hypervaribale regions,Nature 34:877-883(1989); and Al-Lazikani et al., Stand ard Conformations for the canonical stru ctures of immunoglobulins,J.Mol.Biol.273 See 927-948 (1997).

[0103] "Domain antibodies" are immunologically defined as containing only the variable region of the heavy chain or the variable region of the light chain. It is a functional immunoglobulin fragment. In some cases, it contains two or more V H The area is The bivalent domain antibody is covalently bonded by a petit drinker to form a bivalent domain antibody. , 2 V H The region can target the same or different antigens.

[0104] A "bivalent antibody" contains two antigen-binding sites. In some cases, these two binding sites... They have the same antigen specificity. However, bivalent antibodies can be bispecific (see below). I want to be.

[0105] In one embodiment, the monoclonal antibody used herein is a camel monoclonal antibody. This also includes antibodies [e.g., Muyldermans et al., Trends Biochem.]. Sci.26:230(2001);Reichmann et al., J.Immunol.Me thods 231:25(1999);WO 94 / 04678;WO 94 / 255 See 91;US6,005,079. In one embodiment, this invention The first is two V molecules that have been modified to form a single-domain antibody. H Single including domain We provide domain antibodies.

[0106] As used herein, the term "diabody" refers to a small antibody having two antigen-binding sites. This refers to a fragment, and this fragment has a light chain variable domain within the same polypeptide chain. V L ) linked to heavy chain variable domain (VH)(V H -V L or V L -V H ) includes. Using a linker that is too short to allow pairing between two domains on the same strand As a result, those domains are forced to pair with complementary domains of another chain, It generates two antigen-binding sites [EP 0 404 097 B1;WO 93 / 111 61; and Holliger et al., Proc. Natl. Acad. Sci. USA 9 See 0:6444-6448 (1993). Review of manipulated antibody variants and In general, Holliger and Hudson, Nat. Biotechn See ol.23:1126-1136(2005).

[0107] Typically, the antibody or antigen-binding fragment of the present invention expresses its activity on a molar basis. If so, the canine IL-4R (compared to the parent antibody) α at least 1 of the binding activity It possesses 0%. Preferably, the antibody or antigen-binding fragment of the present invention is the parent antibody. Hand Dog IL-4R α At least 20%, 50%, 70%, 80%, 90% of the binding activity, It possesses 95%, 100%, or more. Furthermore, the antibody or antigen binding properties of the present invention Fragments are conserved or non-conserved amino acid substitutions that do not substantially alter their biological activity. It is intended to include (referred to as "conservative variants" or "function-conserving variants" of antibodies) ru.

[0108] "Isolated antibody" refers to a purified state, and in such a state, the molecule is otherwise Biological molecules, such as nucleic acids, proteins, lipids, carbohydrates, or other substances, such as This means that it is substantially free of spore residue and growth medium. Generally, "isolated" The phrase "the complete absence of such a substance, or the absence of water, buffer, or salt" refers to the complete absence of such a substance. This does not mean that they are experimental results of the binding compounds described herein. Or it must not be present in amounts that substantially interfere with therapeutic use.

[0109] As used herein, "chimeric antibody" refers to a variable domain from the first antibody and a variable domain from the second antibody. The antibody has a constant domain, and here, the first antibody and the second antibody are from different species. [US4,816,567; and Morrison et al., Proc. Na [tl.Acad.Sci.USA 81:6851-6855(1984)]. Typically The variable domain is obtained from antibodies ("parent antibodies") from experimental animals such as rodents, and The constitutive domain sequence is obtained from animal target antibodies (e.g., human or dog), and as a result, These chimeric antibodies may induce adverse immune responses in dogs or humans, respectively. However, the antibody levels will likely be lower than those of the parent (e.g., rodents).

[0110] As used herein, the term "canine antibody" refers to canine antibodies and non-canine antibodies (e.g., mouse antibodies). ) This refers to the form of an antibody that contains sequences from both sides of the antibody. Generally, canine antibodies are less It includes at least one, more typically two, variable domains, where, hypervariable All or substantially all of the loop is non-canine immunoglobulin (for example, as illustrated below) As stated, 6 mice anti-canine IL-4Rα Supports CDRs and frames All or substantially all of the work (FR) area (and typically all of the remaining frames) (or substantially all) are canine immunoglobulin sequences. Examples provided herein As described above, canine antibodies, along with canine flame or modified canine flame, are used in mouse anti-canine I L-4R α Contains both 3 heavy chain CDRs and 3 light chain CDRs from the antibody. Canine Anti Further optimizing the effectiveness of the body, for example, canine IL-4R α To strengthen that connection to, Canine IL-4 and / or type I and / or type II IL-4 receptors Or enhances its ability to block the binding of canine IL-13, as illustrated herein. The modified canine flame contains the amino acid changes described above.

[0111] The term "complete canine antibody" refers to an antibody that contains only the canine immunoglobulin protein sequence. Complete canine antibodies are used in mice, in mouse cells, or in mouse cells. When produced in the resulting hybridoma, it may contain mouse carbohydrate chains. Similarly, "mouse antibody" refers to an antibody that contains only mouse immunoglobulin sequences. Complete canine antibodies are found in rats, in rat cells, or derived from rat cells. When produced in a hybridoma, it may contain rat carbohydrate chains. In this context, "rat antibody" refers to an antibody that contains only rat immunoglobulin sequences.

[0112] There are four known IgG heavy chain subtypes of canine IgG, which are IgG-A, Ig These are referred to as GB, IgG-C, and IgG-D. The two known light chain subtypes are lambda It is called Yobikappa.

[0113] The variable region of each light / heavy chain pair forms an antibody binding site. Therefore, generally, intact antibodies It has two binding sites. Except in the case of bifunctional or bispecific antibodies, the two of them The joining sites are generally the same.

[0114] Typically, both the heavy and light chain variable domains are relatively conserved frameworks. Three hypervariable regions located within the FR region, also known as the complementarity determination region (CDR). This includes. CDRs are typically framed to allow binding to specific epitopes. They are aligned by the chromosome region. Generally, the light chain and heavy chain variable chromosomes are aligned from the N-terminus to the C-terminus. Both mains are FR1, CDR1, FR2, CDR2, FR3, CDR3 and FR4 Includes. The assignment of amino acids to each domain is generally done according to Sequences of Prot Ins of Immunological Interest, Kabat et al.; Na tional Institutes of Health,Bethesda,Md. ;5th ed.;NIH Publ.No.91-3242(1991);Kabat Adv. Prot. Chem. 32:1-75 (1978); Kabat et al., J. Bi ol.Chem.252:6609-6616(1977);Chothia et al., JM ol.Biol.196:901-917(1987) or Chothia et al., Nat This is based on the definition in ure 342:878-883 (1989).

[0115] As used herein, the term "hypervariable region" refers to the amino acid residue of an antibody that gives rise to antigen binding. It means base. The hypervariable region is the "complementarity determination region" or "CDR" (i.e., light chain variable). CDRL1, CDRL2, and CDRL3 within the domain, and C within the heavy chain variable domain. [Antibody sequence containing amino acid residues from DRH1, CDRH2, and CDRH3] Kabat et al., Sequences of Proteins, define the CDR region. f Immunological Interest,5th Ed.Public H health service,National Institutes of Hea See Ith, Bethesda, Md. (1991); also, depending on the structure, antibodies Chothia and Lesk, J.Mol.Biol.196 define the CDR area. See 901-917 (1987). The "framework" used in this specification The terms "" or "FR" residues refer to hypervariable residues defined herein as CDR residues. This refers to variable domain residues excluding regional residues.

[0116] In addition to binding and activation of canine immune cells, IL-4R α Canine or canine-like antibodies against The body optimally possesses the following two characteristics: 1. Antibody-dependent cell-mediated cytotoxicity (ADCC) and complement-dependent cell-mediated cytotoxicity (CDC) It lacks effects functionality; and 2. Using industry-standard technologies such as Protein A chromatography-based techniques It can be easily purified on a large scale.

[0117] Not all naturally occurring canine IgG isotypes meet both criteria. No. For example, IgG-B can be purified using protein A, but high levels of A It possesses DCC activity. On the other hand, IgG-A weakly binds to protein A, which is undesirable. It exhibits ADCC activity. Furthermore, IgG-D does not exhibit ADCC activity (IgG-C does, however... IgG-C and IgG-D, both possessing ADCC activity, are derived from protein A. It cannot be purified on the ion. One method of overcoming this problem in this invention is IL-4R α to This is achieved by obtaining specific mutant canine IgG-B antibodies, and such antibodies are A It lacks effects functions like DCC and uses industry-standard protein A chromatography. It can be easily purified using this method.

[0118] "Homologousity" refers to the relationship between two polynucleotide sequences that are optimally aligned. This refers to the sequence similarity between two polypeptide sequences. In both locations, the position is occupied by the same base or amino acid monomer subunit. For example, the positions in each of the two DNA molecules are occupied by adenine. In this case, the molecules are homologous at that position. The percentage of homology is determined by the two sequences. The number of shared homologous positions is divided by the total number of positions being compared, and then multiplied by 100. For example, 6 out of 10 positions in two arrays are optimally aligned. When they are matched or homologous, the two matches The columns are 60% homologous. In general, the comparison is made between two sequences to give maximum homology (%). This is performed when the alignment is correct.

[0119] "Isolated nucleic acid molecule" refers to the case where the isolated polynucleotide is found in nature. Poly(s) that do not contain all or part of a creotide, or which are not naturally linked. DNA linked to nucleotides, genome, mRNA, cDNA or synthetic DNA This refers to RNA, or any combination thereof. For the purposes of this disclosure, specific It should be understood that "nucleic acid molecules containing" the nucleotide sequence do not contain intact chromosomes. Isolated nucleic acid molecules that "contain" the identified nucleic acid sequence are added to the identified sequence. Furthermore, up to 10 or even 20 or more other proteins or parts thereof It may include a coding sequence of fragments, or the nucleic acid sequence listed. It may include functionally linked regulatory sequences that control the expression of the coding region, and It is possible to include / or a vector sequence.

[0120] As used herein, the term "regulatory sequence" refers to a functionally linked sequence in a particular host organism. This refers to the DNA sequence necessary for the expression of the coding sequence. Examples of regulatory sequences suitable for prokaryotes include For example, it includes a promoter, optionally an operator sequence, and a ribosome binding site. Eukaryotic cells use promoters, polyadenylation signals, and enhancers. This is publicly known.

[0121] Nucleic acids can be said to be "functionally linked" if they are functionally linked to another nucleic acid sequence. This is the case when the DNA of the presequence or secretion leader is located in a specific position. It can be said that the peptide is functionally linked to the DNA of the polypeptide. This refers to cases where the protein is expressed as a preprotein involved in secretion, and the promoter or enhancer The reason why a sensor can be said to be functionally linked to a code sequence is that it influences the transcription of that sequence. This is the case when the ribosome binding site is functionally linked to the coding sequence. It can be said that this is true if it is positioned to facilitate translation. Generally speaking, "function" "Continuously linked" means that the linked DNA sequences are continuous and secreted. In the case of a leader, it must be continuous and the leading frame must be consistent. This means that the enhancers do not need to be continuous. The connections are at convenient limiting points. This is achieved by a connection. If such a part does not exist, the usual convention is followed. A synthetic oligonucleotide adapter or linker is used.

[0122] The terms "cell," "cell system," and "cell culture" used herein are interchangeable. And all such expressions include later generations. Therefore, "transformed" and "transformed" The term "transplantation" refers to the primary target cells and their respective cells, regardless of the number of transfers. This includes cultures derived from this. Also, due to intentional or unintentional mutations, all offspring are It is understood that they do not have exactly the same DNA content as the original transformed cells. This also includes offspring of mutants that have the same function or biological activity as those screened. If a different name is intended, it will be clear from the context.

[0123] As used herein, "germline sequence" refers to the sequence of an unreconstructed immunoglobulin DNA sequence. Tastes good. Any suitable unreconstituted immunoglobulin sequence source can be used. Human germline. The sequence is, for example, National Institute of Arthritis and Musculoskeletal and Skin Diseases of the United States National Institutes o JOINSOLVER® germline data on the f Health website It can be obtained from the base. Mouse germline sequences are, for example, Giudicelli et al. [Nu [Described in cleic Acids Res.33:D256-D261(2005)] It can be obtained as described.

[0124] Mouse anti-canine IL-4R α and canine-like mouse anti-canine IL-4R α Characteristics of antibodies This invention relates to isolated mouse anti-canine IL-4R α We provide antibodies and their canine counterparts. Furthermore, the antibody or so in the treatment of diseases, for example, in the treatment of atopic dermatitis in dogs This provides a method for using antigen-binding fragments. In dogs, A, B, C and D There are four IgG heavy chains, which are referred to as [specific terms]. These heavy chains are four different types of canine IgG. These correspond to subclasses, which are referred to as IgGA, IgGB, IgGC, and IgGD. Each of the two heavy chains has one variable domain (VH), as well as CH-1, CH-2 It consists of three constant domains, also called CH-3. The CH-1 domain is a "hinge". Alternatively, it is linked to the CH-2 domain via an amino acid sequence called the "hinge region". ru.

[0125] The DNA and amino acid sequences of these four heavy chains were found by Tang et al. [Vet.Immuno] [First identified by l.Immunopathol.80:259-270(2001)] The amino acid and DNA sequences of these heavy chains were also obtained from the GenBank database. It is available. For example, the amino acid sequence of the IgGA heavy chain is accession number AAL35 301.1 is present, and IgGB has accession number AAL35302.1, IgG C has accession number AAL35303.1, and IgGD has accession number A It has AL35304.1. The canine antibody also has two types of light chains, namely kappa It contains and lambda. The DNA and amino acid sequences of these light chains are from GenBank. It can be obtained from a database. For example, the kappa light chain amino acid sequence is accession number AB. The lambda light chain has accession number ABY 55569.1. To possess.

[0126] In this invention, the amino acid sequences of each of the four canine IgG Fc fragments are The identified boundaries of the CH1 and CH2 domains, as determined by Tang et al. (cited above) Based on: Canine IL-4R α Canine-mutated mouse anti-canine IL-4R binding α Antibodies include mau Anti-dog IL-4R α Along with CDR, canine IgG-A, IgG-B, and IgG-D weights Antibodies containing chains and / or dog kappa light chains are included, but are not limited to these. There is none. Therefore, the present invention is for canine IL-4R α Combined with Type I or Type II IL-4 Isolated anti-canine mice that block the binding of canine IL-4 and canine IL-13 to receptors. IL-4R α and / or canine-like mouse anti-canine IL-4R α Antibodies or their antigenic bonds Provides compatible fragments.

[0127] The present invention further provides a full-length canine heavy chain that can balance with the corresponding light chain to provide canine antibodies. Therefore, the present invention further provides canine mouse anti-canine antigen antibodies (isolated canine mice). Us Anti-Dog IL-4R α It provides antibodies, and also treats diseases, for example in dogs. Methods for using the antibody or its antigen-binding fragment in the treatment of atopic dermatitis provide.

[0128] The present invention also relates to a canine mouse antibody containing a canine crystallizable fragment region (cFc region). Inu IL-4R α The antibody provides, and here, cFc enhances one or more effector functions. They are genetically modified to be reduced or eliminated. In one embodiment of the present invention Genetically modified cFc reduce or eliminate one or more effect functions. In another aspect of the present invention, genetically modified cFc is one or more effectors Enhances function. In one embodiment, a genetically modified cFc region is genetically This is a modified canine IgGB Fc region. In another such embodiment, The genetically modified cFc region is the genetically modified canine IgGC Fc region. In certain embodiments, the effector function is antibody-dependent, which enhances, reduces, or eliminates it. This is cytotoxicity (ADCC). In another embodiment, the effector function increases It is a strong, reduced, or absent complement-dependent cell injury (CDC). Yet another embodiment In this case, the cFc region enhances, reduces, or eliminates both ADCC and CDC. It is genetically modified to achieve this.

[0129] To create a canine IgG mutant lacking effector function, several mutants were used. NuIgGB heavy chains were constructed. These mutants have the following characteristics in the Fc portion of the heavy chain amino acid sequence. This may include one or more of the following single or combined substitutions: P4A, D31A, N63A, G6 4P, T65A, A93G, and P95A. Mutant heavy chains (i.e., such amino acids) Cloning a (substitution-containing) gene into an expression plasmid, and the gene encoding the light chain The plasmid containing the substance was transfected into HEK293 cells. The intact antibody expressed and purified from the cells is used in Fc γ Evaluate the binding of RI and C1q We evaluated their potential for mediating immune effector functions [July 30, 2014] U.S. Provisional Patent Application No. 62 / 030,812, filed on December 16, 2014. U.S. Provisional Patent Application No. 62 / 092,496 (by referring to the full contents of both applications) See (which is incorporated herein).

[0130] This invention also replaces the corresponding natural IgGD hinge region with a hinge region from the following: This product provides modified canine IgGD.

[0131] IgGA: FNECRCTDTPPCPVPEP(SEQ ID NO: 101); IgGB: PKRENGRVPRPPDCPKCPAPEM (SEQ ID NO: 102); or IgGC: AKECECKCNCNNCPCPGCGL (SEQ ID NO: 103).

[0132] Alternatively, the IgGD hinge region can be modified by substituting a serine residue with a proline residue. It can be modified by association: that is, PKESTCKCI P PCPVPES(Sequence ID 104) (Naturally occurring serine residues are found in the proline residues (P) shown in underline and bold.) (More frequently replaced). Such modifications give canine IgGD lacking fab arm replacement. Modified canine IgGD is a standard method of recombinant DNA technology [e.g., Maniati]. s et al., Molecular Cloning, A Laboratory Manual These can be constructed using (1982). To construct these mutants, canine IgGD The nucleic acid encoding the amino acid sequence can be modified to encode modified IgGD. The modified nucleic acid sequence is cloned into an expression plasmid for protein expression.

[0133] Inu IL-4R α The antibody or antigen-binding fragment that binds to is described herein. The complementarity-determining regions (CDRs) 1, 2, 3, 4, 5 or of the listed mouse anti-canine antibody It may contain 6. Such 1, 2, 3, 4, 5, or 6 CDRs are, independently, described below. It can be selected from the CDR sequences of the following. In another embodiment, canine IL-4R α The isolated antibody or its antigen-binding fragment that binds to the mouse light chain CDR- 1. Canine antibody kappa light chain containing CDR-2 and / or CDR-3, and mouse heavy chain CD It contains canine antibody heavy chain IgG, including R-1, CDR-2, and / or CDR-3.

[0134] In other embodiments, the present invention relates to Sequence IDs 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 6 Less than amino acid sequences 6, 67, 68, 69, 70, 71, 72 and / or 73 At least 1- Canine antibody kappa light chain containing 6 different CDRs, and SEQ ID NOs. 74, 75, 76, 77, 7 8, 79, 80, 81, 82, 83, 84, 85, 87, 88, 89, 90, 91, 92 , against the amino acid sequence of 93, 94, 95, 96, 97, 98, 99 and / or 100 and containing at least 80%, 85%, 90%, 95%, 98%, or 99% sequence identity It has canine antibody heavy chain IgG containing 1 to 6 different CDRs, while also having the desired binding and The IL-4R still demonstrates its functional characteristics. α An antibody that specifically binds to or its antigen-binding flag In another embodiment, the antibody or antigen-binding fragrance of the present invention is provided. The component still exhibits the desired bonding and functional properties, while having 0, 1, 2, 3, 4, or 5 Having one or more of the above CDR amino acid sequences containing conserved or non-conserved amino acid substitutions It contains a canine flame containing a combination of kappa light chain and IgG heavy chain sequences.

[0135] Sequence identity means that the amino acids of two polypeptides are optimally aligned. This refers to the degree to which, when aligned, they are the same at the same position. In this specification, the term "identical" to the second amino acid sequence is used to mean that the column is 100% "identical" to the second amino acid sequence. This is the case when the amino acid residues of both sequences are identical. Therefore, the amino acid sequence is second The fact that the amino acid sequences are 50% "identical" to those amino acid sequences is due to the amino acid residues of those amino acid sequences. This is the case when 50% of the base is identical. Sequence comparison is performed on a given protein (for example, comparison). A sequence of amino acid residues contained in a protein or polypeptide (part of the protein) This is done over a certain period. In certain embodiments, selected deletions or insertions are taken into consideration. This could potentially alter the correspondence between two amino acid sequences.

[0136] Sequence similarity includes identical residues and non-identical biochemically related amino acids. Similarity characteristics Biochemically related amino acids that can be shared and exchanged will be considered.

[0137] A "conservatively modified variant" or "conservative substitution" is an amino acid in a protein. However, similar properties (e.g., charge, side chain size, hydrophobic / hydrophilic, backbone conformation) This means that the amino acid is substituted with another amino acid that has properties such as rigidity and stiffness, where the change This can be frequently performed without altering the biological activity of the protein. Generally, polypeptide A single amino acid substitution in a non-essential region of a molecule does not substantially alter its biological activity, as described by those skilled in the art. They are aware of [for example, Watson et al., Molecular Biology of the Gene,The Benjamin / Cummings Pub.Co., See p.224 (4th Ed.; 1987). Also, structural or functional Similar amino acid substitutions are less likely to impair biological activity. Typical conservative substitutions This is shown in Table 1 below. [Table 1] TIFF0007854420000002.tif93145

[0138] Functionally conserved variants of the antibody of the present invention are also included in the present invention. A "specific variant" is one in which desired properties such as antigen affinity and / or specificity are altered. This refers to an antibody or fragment in which one or more amino acid residues have been altered. The eel mutant is created by amino acid substitution with amino acids that have similar characteristics, for example, preserving the characteristics shown in Table 1 above. This includes, but is not limited to, specific amino acid substitutions.

[0139] nucleic acid The present invention further relates to the mouse anti-canine IL-4R disclosed herein. α and / or i Nu-type mouse anti-canine IL-4R α Immunoglobulins of antibodies and their antigen-binding fragments It contains nucleic acids that encode a brin chain (see the examples below).

[0140] Also, the BLAST algorithm (in this case, the parameters of the algorithm are each Selected to give the maximum match between each array over the entire length of the reference array. When a comparison is made using the method described above, the amino acid sequences of the CDR and antibody provided in this invention are In contrast, they are at least approximately 70% identical, preferably at least approximately 80% identical, and more Preferably at least about 90% identical, most preferably at least about 95% (for example) , 95%, 96%, 97%, 98%, 99%, 100%) contain identical amino acid sequences Nucleic acids encoding immunoglobulin polypeptides are also included in the present invention. The present invention further includes BL AST algorithm (in this case, the parameters of the algorithm are the respective reference arrays) (Selected to give the maximum match between each array over the entire length of the ratio) When comparison is performed, at least approximately 70% similarity is observed to any of the reference amino acid sequences. Preferably, at least about 80% similar, more preferably at least about 90% similar. Similar, most preferably at least about 95% (e.g., 95%, 96%, 97%, 9%). Immunoglobulin polypeptides containing similar amino acid sequences (8%, 99%, 100%) It provides nucleic acids that encode this.

[0141] The nucleotide and amino acid sequence identity (%) used herein is defined as C,MacV ector (MacVector, Inc. Cary, NC 27519), Vecto r NTI (Informax, Inc. MD), Oxford Molecular Aligning Group PLC (1996) and Clustal W algorithm Determined using default parameters and default parameters for identity. These commercially available programs may have the same or similar default parameters. It can also be used to determine sequence similarity using a router. Or, for example, default GCG (Genetics Computer Group) using t-parameters Program Manual for the GCG Package, Versi Using a pile-up program on 7, Madison, Wisconsin, Advanced Blast search can be used under the default filter conditions.

[0142] The following references concern the BLAST algorithm, which is often used in sequence analysis. This is the BLAST algorithm: Altschul, SF et al., J.Mol.Bi. ol.215:403-410(1990); Gish, W. et al., Nature Gen et.3:266-272(1993);Madden, TL et al., Meth.Enz ymol.266:131-141(1996);Altschul, SF et al., Nu Cleic Acids Res.25:3389-3402(1997);Zhang , J. et al., Genome Res. 7:649-656 (1997); Wootton, JC et al., Comput. Chem. 17:149-163 (1993); Hanco ck, JM et al., Comput. Appl. Biosci. 10:67-70 (199 4); Alignment Scoring System: Dayhoff, MO et al., “A m evolutionary change in proteins” ,Atlas of Protein Sequence and Structure ,vol.5,suppl.3.MODayhoff(ed.),pp.345-352 (1978);Natl.Biomed.Res.Found.,Washington , DC; Schwartz, RM et al., “Matrices for detecti ng distant relationships”, Atlas of Prote in Sequence and Structure,vol.5,suppl.3( 1978), MODayhoff (ed.), pp.353-358 (1978), Na tl.Biomed.Res.Found.,Washington,DC;Altsc hul, SF, J.Mol.Biol.219:555-565(1991);St ates, DJ et al., Methods 3:66-70 (1991); Henikof f, S. et al., Proc. Natl. Acad. Sci. USA 89:10915-10 919(1992);Altschul, SF et al., J.Mol.Evol.36:2 90-300 (1993); Alignment Statistics: Karlin, S. et al., Proc. N atl.Acad.Sci.USA 87:2264-2268(1990);Karl in, S. et al., Proc. Natl. Acad. Sci. USA 90:5873-58 77(1993); Dembo, A. et al., Ann. Prob. 22:2022-2039 (1994); and Altschul, SF “Evaluating the s statistical significance of multiple dist inct local alignments”, Theoretical and C Computational Methods in Genome Research( S. Suhai (ed.), pp.1-14, Plenum, New York (1997).

[0143] The present invention also provides an expression vector comprising the isolated nucleic acid of the present invention, wherein the nucleic acid This refers to the system that is recognized by the host cell when the host cell is transfected with the vector. It is functionally linked to the sequence. Furthermore, the present invention provides host cells containing the expression vector. Furthermore, a host containing an expression vector encoding an antibody or antigen-binding fragment. Cells are cultured in a culture medium, and an antigen or its antigen-binding fragment is extracted from the host cell or the culture medium. This includes isolating the antibody or its antigen-binding fragment disclosed herein. A method for manufacturing [the product] is provided.

[0144] Epitope binding and binding affinity This invention further relates to mouse anti-canine IL-4R α Similar to antibodies, canine IL-4R α Epitope The present invention provides an antibody or its antigen-binding fragment that binds to an amino acid residue. In terms of administration method, the mouse is anti-canine IL-4R α Antibodies or their antigen-binding fragments This involves the binding of canine IL-4 and canine IL-13 to type I and / or type II IL-4 receptors. It can suppress / block the combination.

[0145] Canine-like mouse anti-canine IL-4R α Antibodies are recombinantly produced by methods known in this field. It may be used as a host for the expression of the antibodies or fragments disclosed herein. The usable mammalian cell lines are well known in this field, including American Type Numerous immortalized cell lines available from Culture Collection (ATCC) These include, among others, Chinese hamster ovary (CHO) cells, NSO SP2 cells, HeLa cells, baby hamster kidney (BHK) cells, monkey kidney cells (COS) ), human hepatocellular carcinoma cells (e.g., Hep G2), A549 cells, 3T3 cells, HEK- It includes 293 cell lines and numerous other cell lines. Mammalian host cells include human, mouse, and rat. This includes dog, monkey, pig, goat, cow, horse, and hamster cells. Particularly preferred Cell lines are selected by determining which cell lines have high expression levels. Other cell lines that can be used include insect cell lines, such as Sf9 cells, amphibian cells, and bacterial cells. Examples include cells, plant cells, and fungal cells. Heavy chains or their antigen-binding portions or flare Recombinant expression vectors encoding the ligation, light chain, and / or its antigen-binding fragments When introducing the antibody into mammalian host cells, the expression of the antibody in the host cells (or More preferably, to enable the secretion of the antibody into the culture medium in which the host cells are cultured. The antibody is produced by culturing the host cells for a certain period of time.

[0146] Antibodies can be recovered from the culture medium using standard protein purification methods. Furthermore, the production process... The expression of the antibody (or other part thereof) of the present invention from the cell line is performed using several known techniques. It can be enhanced by certain conditions. For example, the glutamine synthetase gene expression system (GS system) This is a common approach to enhance expression under certain conditions. The GS system is covered by European Patent No. 0216 Patent Nos. 846, 0256055, and 0323997 and European Patent Applications This is discussed in whole or in part in issue No. 89303964.4.

[0147] Generally, glycoproteins are produced in individual cell lines or transgenic animals. This is characteristic of glycoproteins produced in the cell line or transgenic animals. It will have a glycosylation pattern. Therefore, the individual glycosylation patterns of antibodies The individual cell systems or transgenic animals used to produce the antibody It will depend on the nucleic acid molecule provided in this invention, or All antibodies containing amino acids provided in this invention are capable of having glycosylation patterns. The present invention is independent of n. Similarly, in certain embodiments, non-fucosylated Antibodies with a glycosylation pattern containing only N-glycans may be advantageous because These antibodies, both in vitro and in vivo, respond to their fucosylation This is because it has been shown to typically exhibit greater efficacy than material objects [for example, Shinkaw a et al., J. Biol. Chem. 278:3466-3473 (2003); United States See issues 6,946,292 and 7,214,775.

[0148] The present invention further relates to the mouse anti-canine IL-4R disclosed herein. α Antibody flag It contains ment. The antibody fragment contains the F(ab)2 fragment, which is for example Fab fragments can be produced by enzymatic cleavage of IgG with pepsin. If, it is produced by reduction of F(ab)2 with dithiothreitol or mercaptoethylamine. It can be manufactured. The Fab fragment is V by disulfide crosslinking. H -C H1 Chained ta V L -C L It is a chain. The F(ab)2 fragment now forms two disulfide bridges. These are two more linked Fab fragments. The Fab portion of the F(ab)2 molecule is F is located in between the sulfide bridges. c Includes a portion of the region. F v The fragment is V L Also is V H It is a domain.

[0149] In one embodiment, the antibody or antigen-binding fragment has a heavy chain constant region, For example, the constant region of canine IgG-A, IgG-B, IgG-C, and IgG-D in dogs. The heavy chain constant region or variants thereof are included. In another embodiment, the antibody also The antigen-binding fragment is the light chain constant region, for example, the canine light chain constant region, for example, the lambda region. This includes the kappaine light chain region or its variants. These are not limited examples. Therefore, the canine heavy chain constant region can be derived from IgG-B, and the canine light chain constant region is It is possible that it originates from Pap.

[0150] Antibody modification The present invention provides canine-like mice with anti-canine IL-4R α Antibodies are used, for example, to improve the properties of the antibody. In the canine framework within the variable domain of the parent (i.e., canine) monoclonal antibody It may be manipulated to include modifications to the canine flame residue.

[0151] Experimental and diagnostic applications The present invention provides mouse anti-canine IL-4R α and / or canine-like mouse anti-canine IL-4R α anti The body or its antigen-binding fragments are canine IL-4R α Protein-related diagnostics Assays, e.g., detection of expression related to and / or atopic dermatitis. It can also be useful in this context.

[0152] For example, such a method includes the following steps:

[0153] (a) Mouse anti-canine IL-4R α Antibodies or antigen-binding fragments of an antibody or its substrate (e.g.) For example, a microtiter plate well (for example, the surface of a plastic plate) To do something; (b) Dog IL-4R α Apply the sample to be tested for the presence of the substrate. thing; (c) Wash the plate to remove any unbound material from the sample; (d) IL-4R α Antibodies labeled in a detectable manner that are similarly specific to the antigen (e.g.) For example, applying an enzyme-conjugated antibody; (e) Wash the substrate to remove the unbound labeled antibody; (f) If the labeled antibody is an enzyme conjugate, it is converted into a fluorescent signal by the enzyme. Applying chemical substances; and (g) Detect the presence of a labeled antibody.

[0154] In another embodiment, the labeled antibody is labeled with peroxidase, which is A BTS [e.g., 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfone] It reacts with an acid or 3,3',5,5'-tetramethylbenzidine to produce a detectable color change. This results in... Alternatively, the labeled antibody may contain a detectable radioactive isotope (e.g., 3 H) Marked This can be detected by a scintillation counter in the presence of scintillant. The present invention provides a mouse anti-canine IL-4R α Antibodies can be detected by Western blot or immunoprotein blot. It can be used in lot-based systems.

[0155] Such methods constitute part of the present invention and include, for example, the following:

[0156] (i) Conjugated canine IL-4R α or the film or fragment thereof to be tested in relation to the presence of such fragments Other solid substrates are used in the present invention's mouse anti-canine IL-4R α Antibodies or their antigen-binding fragments To bring into contact with the film. Such films are non-denatured PAGE (polyacrylamide gel electrophoresis). (Action) Gel or SDS-PAGE (Sodium dodecyl sulfate polyacrylamide gel electrolysis) In electrophoresis gel, canine IL-4R α The protein to be tested for the presence of (for example, After electrophoretic separation in the gel, the nitrocellulose that has been transferred... Alternatively, it can take the form of a vinyl-based [e.g., polyvinylidene fluoride (PVDF)] film. The membrane was used to test the mouse anti-canine IL-4R α Before contact with the antibody or its antigen-binding fragment For example, to bind the membrane to nonspecific protein binding sites on the membrane, if desired. Alternatively, you can block it with skim-dried milk or something similar.

[0157] (ii) Wash the membrane once or more to obtain unbound mouse anti-canine IL-4R α antibody or Removal of antigen-binding fragments and other unbound substances; and (iii) Conjugated mouse anti-canine IL-4R α Antibodies or their antigen-binding fragments To detect a

[0158] Detection of bound antibodies or antigen-binding fragments is performed using a secondary antibody labeled in a detectable manner. The body (anti-immunoglobulin antibody) is bound to the antibody or antigen-binding fragment, and the secondary This could be achieved by detecting the presence of an antibody label.

[0159] The mouse anti-canine IL-4R disclosed herein α Antibodies and their antigen-binding flags Menth can also be used in immunohistochemical methods. Such methods constitute part of the present invention. For example, (1) Dog IL-4R α Cells to be tested for the presence of the mouse antimicrobial agent of the present invention NuIL-4R α (2) Contact the antibody with its antigen-binding fragment, and on the cell This includes detecting the antibody or fragment within the cell. The antibody or antigen binding properties If the fragment itself is labeled in a detectable manner, it can be detected directly. Alternatively, the antibody or antigen-binding fragment may be labeled in a detectable manner. It can bind to the secondary antibody that is produced.

[0160] Imaging techniques include SPECT imaging (single-photon emission computed tomography). This includes shadowing or PET (positron emission tomography). Labeling may include, for example, iodine-123. ( 123 I) and technetium-99m( 99m Tc) (For example, SPECT image) (something that is combined with ing), or 11 C, 13 N, 15 O or 18 F (for example, PET (Those combined with imaging), or those containing indium-111 [e.g., Go rdon et al., International Rev. Neurobiol. 67:385 See -440 (2005).

[0161] Cross-blocking antibodies Furthermore, the present invention's anti-canine IL-4R α Antibodies and their antigen-binding fragments are, The antibodies and fragments described in the document bind to the same canine IL-4R α to Any antibody or antigen-binding fragment that binds to the epitope in, and NuIL-4R α Regarding binding, the antibodies or fragments described herein (partially or (Completely) cross-blocked, or (partially) blocked by the antibody or fragment described herein. Any antibody or antigen-binding fragment that is cross-blocked (either partially or completely), as well as This includes any of their variants.

[0162] The cross-blocking antibodies and their antigen-binding fragments described herein are (later The antibodies disclosed herein (based on the CDR described in Example 5), i.e. , 1A3, 1A9, 1B12, 10C12, 10F2, 10E10, 10G8 and / or or 11D3, or more specifically 11B6 and / or 6C12, and more specifically Layer details include their capabilities that cross-compete with 4D8, 4H3, 2E2, and / or 11H2. Based on this, a standard binding assay (e.g., BIACore®, exemplified below) is used. It can be identified in ELISA or flow cytometry. For example, A standard ELISA assay is available, in which case recombinant canine IL-4R α Tan The protein is immobilized on a plate, one of the antibodies is fluorescently labeled, and the binding of the labeled antibody competes with that of the other antibodies. Evaluate the ability of the unlabeled antibody. Additionally, evaluate the ability of the antibodies to cross-compete, either as an alternative or by substitution. BIAcore® may be used for this purpose. For example, 1A3, 1A9, 1B 12, 10C12, 10F2, 10E10, 10G8 and / or 11D3, or For more details see 11B6 and / or 6C12, and even more details see 4D8, 4H 3, 2E2 and / or 11H2, Canine IL-4R α The test antibody inhibits binding to it. This means that the test antibody is 1A3, 1A9, 1B12, 10C12, 10F2, 10E1 0, 10G8, 11D3, 11B6, 6C12, 4D8, 4H3, 2E2 and / or 11H2 and Inu IL-4R α It is possible to compete with respect to joining to, and therefore, in some cases , 1A3, 1A9, 1B12, 10C12, 10F2, 10E10, 10G8, 1 Same as 1D3, 11B6, 6C12, 4D8, 4H3, 2E2 and / or 11H2, Inu IL-4R α This indicates that it can bind to the epitope above. As described above, the anti- NuIL-4R α An antibody that binds to the same epitope as either the antibody or the fragment. The fragment also constitutes a part of the present invention.

[0163] Pharmaceutical composition and administration Canine-like mouse anti-canine IL-4R α Pharmaceutical compositions of antibodies or their antigen-binding fragments Alternatively, to produce a sterile composition, it may be mixed with a pharmaceutically acceptable carrier or excipient. [For example, Remington's Pharmaceutical Sciences] es and US Pharmacopeia:National Formula ry, Mack Publishing Company, Easton, PA (198 See 4).

[0164] Formulations of therapeutic and diagnostic substances include, for example, lyophilized powders, slurries, aqueous solutions, or It may be manufactured in the form of a suspension by mixing it with an acceptable carrier, excipient, or stabilizer. [For example, Hardman et al., (2001) Goodman and Gilman's] s The Pharmacological Basis of Therapeut ics, McGraw-Hill, New York, NY; Gennaro (2000 )Remington:The Science and Practice of P Harmacy, Lippincott, Williams and Wilkins, Ne w York, NY; Avis et al. (eds.) (1993) Pharmaceutical Dosage Forms:Parental Medications,Marc el Dekker, NY; Lieberman et al. (eds.) (1990) Pharmace utical Dosage Forms:Tablets,Marcel Dekke r, NY; Lieberman et al., (eds.) (1990) Pharmaceutical Dosage Forms:Disperse Systems,Marcel Dek ker, NY; Weiner and Kotkoskie (2000) Excipient Toxicity and Safety,Marcel Dekker,Inc. [See New York, NY]. In one embodiment, the anti-IL of the present invention -4R α Dilute the antibody to an appropriate concentration in sodium acetate solution (pH 5-6), and then add NaCl. Alternatively, sucrose may be added for isotonicity. Additional substances may be added to enhance stability, for example. Polysorbate 20 or polysorbate 80 may be added.

[0165] The toxicity and therapeutic efficacy of the antibody composition, administered alone or in combination with another therapeutic agent, are, for example, For example, LD 50 (A dose that is lethal to 50% of the population) and ED 50 (50% of the group smell Standard measurements in cell cultures or experimental animals to determine the therapeutically effective dose. It can be determined by various pharmaceutical methods. The dose ratio of toxic effect to therapeutic effect is the therapeutic coefficient (LD50). 50 / ED 50 ) In certain embodiments, antibodies exhibiting a high therapeutic coefficient are desirable. The data obtained from their cell culture assays and animal studies are for use in dogs. It can be used in determining the range of dosage. The dosage of such compounds is preferably, ED with little to no toxicity 50 The circulating concentration is within the range of [the specified amount]. This range may vary depending on the dosage form and route of administration used.

[0166] The mode of administration can vary. Suitable routes of administration include oral, rectal, transmucosal, intestinal, and parenteral administration. Intramuscular, subcutaneous, intradermal, intramedullary, intrasacral, direct ventricle, intravenous, intraperitoneal, intranasal, intraocular, inhalation, This includes blowing, local, cutaneous, transdermal, or intra-arterial delivery. In certain embodiments, mouse antibodies Inu IL-4R α Antibodies or their antigen-binding fragments are administered via invasive routes such as injection. It can be administered. In another embodiment of the present invention, mouse anti-canine IL-4R α Antibodies The antigen-binding fragment or its pharmaceutical composition is administered intravenously, subcutaneously, intramuscularly, or arterially. It is administered by or inhalation, aerosol delivery, or via non-invasive routes (e.g., orally, for example). Pills, capsules, or tablets are also within the scope of the present invention.

[0167] The composition may be administered using medical devices known in the art. For example, the present invention The pharmaceutical composition is administered via a subcutaneous injection needle (for example, a pre-filled syringe or an automated injection device). It may be administered by injection. The pharmaceutical compositions disclosed herein are, for example, U.S. patents. No. 6,620,135, No. 6,096,002, No. 5,399,163, No. 5,3 No. 83,851, No. 5,312,335, No. 5,064,413, No. 4,941,8 Apparatus disclosed in Patent No. 80, No. 4,790,824 or No. 4,596,556 It can also be administered using needleless subcutaneous injection devices.

[0168] The pharmaceutical compositions disclosed herein may also be administered by injection. Examples of well-known implants and modules for this purpose include U.S. Patent No. 4,48 No. 7,603 (This is a portable micro-injection pump for dispensing medicine at a controlled rate) (Disclosure of the patent), U.S. Patent No. 4,447,233 (This is a pharmaceutical patent that allows for precise injection speed.) U.S. Patent No. 4,447,224 (discloses a pharmaceutical infusion pump for transporting pharmaceuticals) This discloses a variable flow rate implantable infusion device for continuous drug delivery, U.S. Patent Patent No. 4,439,196 (This refers to a penetrating drug having a multi-chamber compartment) This includes those disclosed in (disclosing a material handling system). Numerous other such in The plant, transport systems, and modules are well known to those skilled in the art.

[0169] Alternatively, mouse anti-canine or canine-modified mouse anti-canine IL-4R α Antibodies are not systemic Locally, for example, often by depot or sustained-release formulations, the condition is characterized by immunopathology. The antibody is administered by direct injection into the affected arthritis joint or pathogen-induced lesion. Furthermore, targeted drug delivery systems can be used to characterize, for example, immunopathologically. Lipids coated with tissue-specific antibodies that target arthritis joints or pathogen-induced lesions. The antibody can be administered within the liposome. The liposome is targeted to the affected tissue. It will likely be selectively taken up by the affected tissue.

[0170] Administration is based on the serum or tissue turnover rate of the therapeutic antibody, the severity of symptoms, and the immunogen of the therapeutic antibody. Several factors, including sex and the accessibility of target cells in the biological matrix, can be left to the left. It is corrected. Preferably, the administration regimen is sufficient to bring about improvement in the target condition. While administering therapeutic antibodies, undesirable side effects are minimized at the same time. The amount of biological material transported is, in part, individual therapeutic antibodies and the condition being treated. It depends on the severity. Guidelines are available for selecting an appropriate dose of therapeutic antibody [e.g.] For example, Wawrzynczak, Antibody Therapy, Bios Sci. entific Pub.Ltd,Oxfordshire,UK(1996);Kre sina (ed.) Monoclonal Antibodies, Cytokines a nd Arthritis,Marcel Dekker,New York,NY(1 991); Bach (ed.) Monoclonal Antibodies and Pe. ptide therapy in Autoimmune Diseases,Mar cel Dekker, New York, NY (1993); Baert et al., New Engl.J.Med.348:601-608(2003);Milgrom et al.,Ne w Engl.J.Med.341:1966-1973(1999);Slamon et al. ,New Engl.J.Med.344:783-792(2001);Beniam inovitz et al., New Engl. J. Med. 342:613-619 (2000 );Ghosh et al., New Engl.J.Med.348:24-32(2003); Lipsky et al., New Engl. J. Med. 343:1594-1602 (200 See 0).

[0171] Determining the appropriate dosage is, for example, known in the art to affect treatment. This is done by a veterinarian using the suspected parameter or factor. Generally, the dosage is Start with a dose somewhat lower than the optimal dose, and then compare the desired or optimal efficacy with the negative side effects. Gradually increase the dosage until the desired effect is achieved. Important diagnostic measures include, for example, measures of inflammation symptoms. This may include the levels of inflammatory cytokines produced.

[0172] The antibodies or antigen-binding fragments disclosed herein can be injected by continuous injection. Or for example, daily, 1-7 days a week, weekly, bi-weekly, monthly, bi-monthly, 4 times a year, 2 times a year, once a year. It can be administered at any interval. Administration methods include, for example, intravenous, subcutaneous, topical, oral, and nasal. It can be administered intracavitary, rectally, intramuscularly, intracerebrally, intraspinally, or by inhalation. The total weekly dose is Generally, at least 0.05 μg / kg body weight, and more generally, at least 0.2 μg / kg , 0.5μg / kg, 1μg / kg, 10μg / kg, 100μg / kg, 0.25mg / kg, 1.0mg / kg, 2.0mg / kg, 5.0mg / kg, 10mg / kg, 2 5 mg / kg, 50 mg / kg or more [e.g., Yang et al., New Engl.J] .Med.349:427-434(2003); Herold et al., New Engl. J.Med.346:1692-1698(2002);Liu et al., J.Neurol. Neurosurg.Psych.67:451-456(1999);Portiel ji et al., Cancer Immunol. Immunother. 52:133-144 See (2003). Also, the canine-like mouse anti-canine IL-4R in the serum of the subjects. α The predetermined target concentrations of the antibody, for example, 0.1, 0.3, 1, 3, 10, 30, 100, Administration may be carried out to achieve a concentration of 300 μg / ml or higher. In other embodiments, Invention of canine-like mouse anti-canine IL-4R α Antibodies are administered subcutaneously or intravenously, weekly, bi-weekly, or every four weeks. "Every other time", monthly, every other month, or four times a year, 10, 20, 50, 80, 100, 200, 50 It is administered at doses of 0, 1000, or 2500 mg per subject.

[0173] Furthermore, the binding of canine IL-4 and canine IL-13 to type I and type II IL-4 receptors. Anti-canine IL-4R vaccine is used to produce antibodies that block it. α mAb Recognized antigen peptides may be used. Such vaccines are used for atopic dermatitis. These antigen peptides may be useful as a therapeutic vaccine for mites. To use as a peptide, one or more of these peptides are chemically or recombinant DNA technology By binding these peptides to other carrier proteins, the immunogenicity of these peptides is enhanced. It is possible to generate specific antibodies. This is because the peptide is bound to the carrier protein. The technology is known to those skilled in the art. For vaccinating animals, peptide vaccines are used. Used to vaccinate animals via M, S / C, oral, spray, or intraocular routes. Peptide vaccines can be derived from bacteria, viruses, yeast, or baculoviruses. It can be used as a subunit protein. Alternatively, such a peptide vaccine Chin can perform such peptide vaccines as can be done by methods known to those skilled in the art. The peptide can be transported after administration of various viral or bacterial vectors that express the vector. The vaccine can be administered in doses of 1 to 1000 μg, and an adjuvant may be added if desired. It may contain permissible pharmaceutical carriers.

[0174] As used herein, “suppress,” “treat,” or “treat” refers to a disorder. This includes delaying the onset of symptoms and / or reducing the severity of symptoms of such disorders. Furthermore, it improves existing uncontrolled or undesirable symptoms, prevents additional symptoms, and This includes improving or preventing the underlying cause of such symptoms. Therefore, the term also includes disorders, diseases, and The target is vertebrates that have symptoms, or those that may develop such disorders, diseases, or symptoms. This demonstrates beneficial results for vertebrates with potential.

[0175] The terms "therapeutic effective dose," "therapeutic effective amount," and "effective dose" as used herein are defined as follows: When administered alone or in combination with additional therapeutic agents to cells, tissues, or subjects, it can cause disease Alternatively, one or more symptoms of the disease or condition, or measurable progression of such disease or condition. The present invention's canine-like mouse anti-canine IL-4R is effective in bringing about significant improvement. α Antibodies This refers to the amount of the antigen-binding fragment. The therapeutically effective dose is further defined as the amount of at least the symptoms. Partial improvement, such as treatment, cure, prevention or improvement of the related medical condition, or such The binding compound is sufficient to bring about an increased rate of treatment, cure, prevention, or improvement of the condition. It refers to the quantity. The therapeutically effective dose is when applied to individual active ingredients administered alone. This pertains only to that component. The therapeutically effective dose, when applied to a combination, Regardless of whether it is administered in combination, sequential, or simultaneous doses, the therapeutic effect This refers to the combined amount of active ingredients that are produced. The effective amount of a therapeutic substance is at least 10 %, usually at least 20%, preferably at least about 30%, more preferably less The improvement in the diagnostic scale or parameter is 40%, most preferably at least 50%. The effective dose is also used when subjective scales are used to assess the severity of the disease. This can lead to improvements in subjective scales.

[0176] Other combination therapies As already described, the present invention provides canine-like mice with anti-canine IL-4R α Antibodies or so The antigen-binding fragment and / or antigen peptide is one or more other therapeutic substances (e.g.) (The inhibitors described in the next paragraph) and / or mice (or canine mice) ) Can be co-administered with anti-canine TSLP antibodies [see US8,791,242]. The antibody can be bound to the substance (as an immune complex), and / or The substance or other antibodies may be administered separately. In the latter case (separate administration), the antibodies The substance can be administered before, after, or simultaneously with other known therapies, or it can be administered in combination with other known therapies.

[0177] kit Furthermore, pharmaceutically acceptable carriers and / or inhibitors, such as Janus ) kinase (JAK) inhibitors, such as oclacitinib [See WO 2013 / 040241], spleen tyrosine kinase (SYK) Inhibitor [see, for example, US8,759,366], or TH2 details Antagonists for chemoattractant receptor homolog molecules expressed on cells [e.g., W O 2010 / 099039; WO 2010 / 031183; and US8,54 See 6,422] (but not limited to these) one or more additional Along with the ingredients, IL-4R α The antibodies or antigens described herein that specifically bind to the specified antibody or antigen. Binding fragments (e.g., canine mouse anti-canine IL-4R) α Antibodies or their antigen binding A kit containing one or more components including (but not limited to) sex fragments. Provides the binding compound and / or inhibitor described immediately beforehand, pure It may be formulated as a composition or in combination with a pharmaceutically acceptable carrier in a pharmaceutical composition. ru.

[0178] In one embodiment, the kit contains the binding composition of the present invention (e.g., canine mau). Anti-dog IL-4R α The antibody or its pharmaceutical composition) in one container (for example, sterile glass) (or plastic vial) and the pharmaceutical composition and / or the inhibitor Place the tar in another container (for example, a sterile glass or plastic vial).

[0179] If the kit contains a pharmaceutical composition for parenteral administration to a subject, the kit shall be such The kit may include a device for administering the medication. For example, the kit may include one or more subcutaneous injection needles or This may include other injection devices. The kit also includes pharmaceutical compositions and dosage forms in the kit. This may include an attached leaflet containing information about the enclosed medicine. Generally, such information is included in the enclosed medicine Assisting pet owners and veterinarians in the effective and safe use of compositions and dosage forms. For example, the following information regarding the combination of the present invention is provided in the accompanying document (insert). Information may be provided on: pharmacokinetics, pharmacodynamics, clinical trials, efficacy parameters, indications and dosage, and contraindications. Warnings, precautions, adverse reactions, overdose, appropriate dosage and administration, supply methods, proper storage Conditions, reference materials, manufacturer / distributor information, and patent information.

[0180] For convenience, the antibodies or specific binding substances disclosed herein are provided as a kit. In other words, a package containing a predetermined amount of reagents with instructions for performing a diagnostic or detection assay. It may be provided as a compounded combination. If the antibody is enzyme-labeled, the kit The enzyme requires substrates and cofactors (e.g., detectable chromophores or fluorescence). It contains substrate precursors that give aggregates. It also contains other additives, such as stabilizers and buffers (e.g., This may include blocking buffer or cell lysis buffer, etc. Various reagents The relative volume is large enough to result in a solution concentration of the reagent that substantially optimizes the sensitivity of the assay. It can vary. In particular, the reagent provides an excipient when a reagent solution of an appropriate concentration is dissolved. It may be supplied as a dry powder, usually freeze-dried powder. [Brief explanation of the drawing]

[0181] [Figure 1] Figure 1 shows the reactivity of purified mouse anti-canine IL-4Rα monoclonal antibodies (mAbs) to the extracellular domain of canine IL-4Rα. Various mouse mAbs were tested by ELISA for their binding to the extracellular domain of canine IL-4Rα. The tested mAbs have the following names. [ka]

[0182] The horizontal axis represents the log concentration of the added mAb, and the vertical axis represents the optical concentration obtained by ELISA. This indicates. [Figure 2] Figure 2A shows the dose-response curve for the binding of canine IL-4 to canine IL-4Rα expressed on the surface of CHO cells using a cell-based CHO-cIL-4Rα binding assay. The horizontal axis represents the log concentration of added IL-4, and the vertical axis represents the mean fluorescence intensity (MFI) using FACS. Figure 2B shows the dose-response curve for CHO-cIL-4Rα using the following mouse anti-canine IL-4Rα monoclonal antibody (mAb). [ka]

[0183] The horizontal axis shows the log concentration (nM) of the added mAb, and the vertical axis shows the average concentration when using FACS. The uniform fluorescence intensity (MFI) is shown. The maximum effective concentration (EC50) for each antibody is also shown. This is shown in Table 2 below. [Figure 3] Figures 3A and 3B show the results of serially diluted individual mouse anti-canine IL-4Rα monoclonal antibodies (mAbs) regarding the binding of IL-4 to cell-based CHO-cIL-4Rα. Figure 3A shows monoclonal antibodies that individually block the binding of IL-4 to cell-based CHO-cIL-4Rα. [ka]

[0184] This shows the concentration-dependent capacity. Figure 3B shows the cell-based CHO-cIL-4R α IL-4 Monoclonal antibodies that individually block the binding of these antibodies. [ka]

[0185] This shows the concentration-dependent capability. The horizontal axis represents the log concentration (nM) of the added mAb, and the vertical axis represents, This shows the average fluorescence intensity (MFI) when using FACS. [Figure 4] Figure 4 shows the binding of chimeric and canine monoclonal antibodies to canine IL-4Rα as evaluated by ELISA. The dose-dependent reactivity of canine monoclonal antibodies to the canine IL-4 receptor alpha chain is as follows: [ka]

[0186] Examples Example 1 stomach Identification and cloning of the IL-4 receptor α chain receptor The cDNA encoding the estimated full-length canine IL-4 receptor alpha chain (SEQ ID NO: 1) was used in Ge nbank database (accession number XM) 547077.4; US 7,20 Identified by a search of 8,579 B2 (see also). This presumed cDNA is 25A It codes for 823 amino acids (SEQ ID NO: 2), including the leader sequence of amino acids, and access Edition number XP Identified as 547077.3. Mature canine IL-4 receptor α chain. The protein (SEQ ID NO: 4) is the human IL-4 receptor α chain (accession number NP). 000 409.1) showed 65% identity and porcine IL-4 receptor α chain (accession number) No. NP Shares 70% identity with 999505.1). Estimated mature canine IL-4 The receptor α-chain protein is coded by the nucleotide sequence identified as SEQ ID NO: 3. The comparison between the presumed mature IL-4 receptor α chain and the known sequence of the human IL-4 receptor α chain is as follows: We identified the extracellular domain (ECD) of the mature canine IL-4 receptor α-chain protein. This is referred to as Sequence ID 6. Mature dogs IL-4 The DNA sequence encoding the ECD of the receptor α chain has been identified as Sequence ID No. 5. [ka] TIFF0007854420000009.tif212157TIFF0007854420000010.tif199157TIFF0007854420000011.tif71156

[0187] Example 2 Mouse anti-canine IL-4 receptor alpha chain antibody Production of anti-canine IL-4 receptor α-chain monoclonal antibody : A total of 3 Balc / c mice were given multiple doses (10 μg each) over a 17-day period. The cells were immunized. The immunizing antigen was canine IL-4R alpha chain extracellular domain (ECD)-human F It was a c-fusion protein. After immunization, serum was collected from each mouse and the canine IL-4 receptor was analyzed. Reactivity to rufa chain ECD HIS-tagged proteins was tested in the best serum. Splenocytes of mice with anti-IL-4 receptor alpha chain ECD titer are used to create myeloma P3X63A The cells were fused with the g8.653 cell line. Approximately two weeks after fusion, the superior cells of the presumed hybridoma cells were observed. Their response to IL-4 receptor alpha chain ECD HIS-tagged protein Sex was tested by ELISA. The strong positive signal shown in ELISA was observed in H Ibridomas were subcloned by limiting dilution, and canine IL-4 receptor alpha chain EC was obtained. The reactivity to D HIS-tagged proteins was tested again.

[0188] Confirmation of the reactivity of monoclonal antibodies against the canine IL-4 receptor α chain. : Anti-canine IL-4 receptor alpha chain ECD secreted by hybridoma The body's reactivity was confirmed by ELISA. CELLine bioreactor (Integ Hybridoma cells were cultured for 10-30 days using ra-biosciences. Initially, 4 mM L-glutamine and 10% Ultra Low IgG bovine fetus Cells were maintained in DMEM supplemented with infant serum (FBS) (Gibco). In the bioreactor cell chamber, the same medium with an increased FBS concentration of 20% Approximately 2 x 10⁶ hybridoma cells in 15 mL 6 Seeded at a cell density of cells / mL. The chamber contains 1 L of nutrient medium (4 mL L-glutamine and 2% standard FBS). The cell chamber was filled with DMEM. The hybridoma in the cell chamber was incubated for approximately 3 to 7 days. 2.5 × 10 7 The cells were grown to a certain level (cells / mL). Then, 10 mL of cell suspension was added to the cell channel. The cells were collected from the bars and replaced with fresh culture medium to allow for cell regrowth and subsequent harvesting. Repeat this procedure as needed to obtain a suitable amount of mAb from the ibridomatous clone. The recovered cell suspension was centrifuged, and the supernatant was filtered through a 0.2 micron filtration membrane. Antibodies For purification, Protein G Sepharose 4 First Flow epharose 4 Fast flow) 5mL column (GE Healthcar e) The supernatant of each clone was purified by gravity flow. Tris-EDTA(T E) After washing with buffer (pH 8.0), wash with 0.1M glycine buffer (pH 2.0). 7) Elute the bound antibody using the solution, then use 1M Tris (pH 8.0) to adjust the pH. Neutralization was performed. Centriprep YM-10kDa NMWL centrifugal filter The antibody was concentrated using a Millipore nib, and the buffer was converted to phosphate-buffered sodium chloride. The solution was replaced with water (PBS). Antibody concentration was quantified by spectrophotometric analysis. Purified anti-canine I L-4 receptor α chain mAb with HIS tagged ECD domain of canine IL-4 receptor alpha chain The reactivity to the substance was tested by ELISA as follows: HIS-tagged substance Nu IL-4 receptor alpha chain protein coated with a coating buffer (carbonate / bicarbonate) Dilute to 10 μg / mL in pH 9.0 and plate in a 96-well flat-bottom ELISA plate (NU Dispense 100 μl / well into the NC (Non-Cellulose) plate. Incubate the plate at 4°C overnight. Next, the plate is subjected to a phosphate solution containing 0.05% Tween-20 (PBST). Wash three times with saline solution. Then add 200 μl of blocking buffer (5% of PBST). Add skim milk to each well and incubate the plate at 30°C for 60 minutes. Then wash the plate three times with PBST. Next, diluted in blocking buffer Add 100 μl of the test mAb to the first well of a suitable row. Then add the test mAb to a suitable plate. Dilute 3-fold to the plate position. Incubate the plate at 37°C for 60 minutes, then... Wash the lats three times with PBST. Then, fill with 100 μl / well of 1:2,000 dilution. Add Sladish peroxidase-conjugated goat anti-mouse IgG (KPL) to the plate. Next, incubate this at 37°C for 60 minutes. Then, sterilize the plate with PBST for 3 minutes. Wash once, then add 100 μl / well of 3,3',5,5' tetramethylbenzidine (TMB Add the substrate (obtained from KPL) to the plate. Allow the color reaction to proceed at 37°C for 5-20 minutes. Afterward, measure the absorbance at 650 nm.

[0189] Various mouse anti-canine IL-4R α Monoclonal antibodies (mAbs) are used in canine IL-4R α Their ability to bind to the extracellular domain was assayed by ELISA. Figure As shown in 1, the majority of these mAbs exhibit positive (positive) dose-dependent binding.

[0190] Example 3 Heavy and light chain variable domains of anti-canine IL-4 receptor alpha chain monoclonal antibody Identification of DNA and putative protein sequences After isolating mRNA from each hybridoma using standard molecular biological methods, Identify the DNA sequences of the VH and VL strands from these hybridomas. The sequence numbers for the VL DNA and putative amino acid sequence are shown below. The signal sequence codes are... The DNA and amino acids corresponding to the putative signal sequence are underlined, and for CDR Corresponding items are shown in bold, while FR is neither underlined nor bolded (i.e., C Gnar array -FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4). [ka] TIFF0007854420000013.tif212157TIFF0007854420000014.tif199156TIFF0007854420000015.tif205156TIFF0007854420 000016.tif216157TIFF0007854420000017.tif209157TIFF0007854420000018.tif211157TIFF0007854420000019.tif15152

[0191] Example 4 Construction of a CHO cell line expressing the canine IL-4 receptor alpha chain and ligand blockade Use in Sei Full-length canine IL-4 receptor alpha chain (cIL-4R α ;codes sequence number 4) The gene was synthesized and subcloned into a mammalian expression vector. The resulting plasmid was then converted to C HO DG44 cells were transfected. 48 hours after transfection, Cells were diluted in a 96-well plate to obtain single-cell clones. After 4 weeks of incubation... After the experiment, approximately 130 clones were obtained. Anti-cIL-4Rα monoclonal antibody 6B2 was used. By using FACS, all of the clones regarding the expression of cIL-4Rα can be screened. Three clones were selected for stability evaluation. FACS was used to determine the 20th passage. Stability was monitored over the period.

[0192] The binding of canine IL-4 to canine IL-4R alpha expressed on the surface of CHO cells To evaluate the ability of a monoclonal antibody specific to nuIL-4 receptor alpha to block it. The ligand blockade assay was set up as follows.

[0193] Reagents and equipment: • Cell growth medium: CD OptiCHO medium + 8mM L-glutamine + 0 0.018% F-68; • FACS buffer: BD Pharmingen stain buffer (BD catalog number 554657); · R-phycoerythin-bound streptavidin (Li fe Technologies: SB66); • Inu IL-4 (R&D system, catalog number 754-CL / CF); • Light used to biotinylate canine IL-4 according to the manufacturer's recommendations. ning-Link Biotin Binding Kit Type A (Novus: 704-0010) • Flow cytometer: BD Accuri-C6.

[0194] procedure: 1. CHO-DH44-canIL-4Rα cells with a viability of 96% or more, 2-4 × 1 0 6 The cells were grown to a level of [number] cells / mL.

[0195] 2. Centrifuge the cells to allow them to settle, discard the supernatant, and divide the cells into 2 × 10⁶ cells. 7 FAC up to cells / mL It was suspended in S buffer.

[0196] 3. The cells were dispensed into a U-shaped 96-well plate (50 μl per well).

[0197] 4. Anti-canine IL-4Rα mAb in FACS buffer on a 96-well plate. The solution was diluted 3-fold from 50 μg / mL in the upper wells to the lower wells.

[0198] Transfer 5.50 μl of each dilution (Ab) into a cell plate, then incubate on ice for 30 minutes. I did it.

[0199] 6. The cells were washed twice with FACS buffer.

[0200] 7. Cells are treated with 0.32 μg / mL of biotinylated canine IL-4 in FACS buffer. The solution was resuspended in 100 μl and incubated on ice for 30 minutes.

[0201] 8. The cells were washed twice with FACS buffer.

[0202] 9. Cells in R-phycoerythrin-bound streptavidin (1) in FACS buffer. The solution was resuspended in 100 μl of a 1000 dilution and incubated on ice for 30 minutes.

[0203] 10. The cells were washed twice with FACS buffer.

[0204] 11. The cells were placed in 300 μl of FACS buffer.

[0205] 12. BD Accuri-C6 reads 10,000 cells per sample. I got it.

[0206] 13. The obtained readouts are analyzed using FlowJo to determine the average fluorescence intensity. It achieved a degree (MFI).

[0207] Canine IL-4R expressed on the surface of CHO cells α Uses regarding the binding of canine IL-4 to Dose-response curves based on cells, CHO-cIL-4R α (Figure 2A) (See reference). The maximum half-liquid effective concentration (EC50) of 25 nM was determined from this curve. Next, mouse anti-canine IL-4R α Monoclonal antibodies (mAb): 11B6, 4D8, CHO-cIL-4R with 4H3, 2E2, 11H2 and 6C12 α Regarding the joining Dose-response curves were obtained (see Figure 2B). The maximum half dose effective for each antibody was determined. The concentrations (EC50) are shown in Table 2 below. [Table 2]

[0208] Next, mouse anti-canine IL-4R α Monoclonal antibodies (mAbs) are used in cell-based CH O-cIL-4R α Assays were conducted regarding their ability to block the binding of canine IL-4 to [the target]. As shown in Figure 3A, there were five mAbs, namely 11B6, 4D8, 4H3 2E2 and 11H2 showed significant blocking ability. In a supplementary study, a sixth mAb was used. Tested (6C12) and compared with one of the five test mAbs (11H2) (Figure 3A). As is clear from Figure 3B and Table 2, 6C12 mAb was significantly more effective than 11H2 mAb. It has a high maximum half-dose inhibitory concentration (IC50). In Figures 3A and 3B and Table 2, As can be acknowledged, anti-cIL-4R α Monoclonal antibodies 4D8, 2E2, 4D8 and The four components of 11H2 showed excellent blocking ability.

[0209] Example 5 Amino acid sequence of mouse CDR [ka] TIFF0007854420000022.tif179154TIFF0007854420000023.tif123155 [Table 3] TIFF0007854420000025.tif40146

[0210] Example 6 Epitope mapping of mouse anti-canine IL-4 receptor alpha antibody The interaction of the antibody with the corresponding cognitive protein antigen is due to the specific amino acids (parato) of the antibody. The process is mediated by the binding of an epitope to a specific amino acid (epitope) of the target antigen. Epitopes are antigenic determinants that trigger a specific response by immunoglobulins. It consists of a group of amino acids on the surface of an antigen. The protein of interest can be identified by various antibodies. It may contain several epitopes that are recognized by antibodies. Epitope recognized by antibodies are linear epitopes. They are classified as pitopes or conformational epitopes. Linear epitopes are tan It is formed by a series of continuous sequences of amino acids in a protein, while conformational proteins Pitopes are discontinuous in the primary amino acid sequence (for example, they are far apart). It is composed of amino acids that combine during three-dimensional protein folding.

[0211] Epitope mapping is the amino acid sequence on the target antigen that is recognized by the antibody. This refers to a method for identifying an epitope. Monoclonal antibodies (mA) on the target antigen. Identifying epitopes recognized by b) has important applications. For example, it can be used in novel therapies. It can assist in the development of drugs, diagnostic agents, and vaccines. Epitope mapping can also optimize treatment. It can assist in the selection of therapeutic mAbs and help elucidate their mechanisms of action. Furthermore, it can aid in IL-4 receptors. Information on epitopes related to body alpha reveals unique epitopes and provides vaccine protection. It may reveal the effects or pathogenicity. Epitope identification also involves carrier proteins. This involves the chemical or genetic coupling of identified peptide epitopes to other immunostimulants. This could lead to the development of subunit vaccines based on this.

[0212] Epitope mapping can be performed using polyclonal or monoclonal antibodies. It is possible, and depending on the expected properties of the epitope (i.e., linear pair conformation) Several methods are used to identify epitopes. Mapping of linear epitopes... The approach is more direct and relatively easier to implement. For this purpose, linear epi Commercial services for tope mapping often use peptide scanning. In this case, a set of duplicated short peptide sequences of the target protein is chemically synthesized, and the desired result is obtained. We will test their ability to bind to antibodies. This method is rapid and high-throughput. It is relatively inexpensive to perform. On the other hand, mapping discontinuous epitopes is technically more This is difficult and requires more specialized techniques, such as monoclonal antibodies against corresponding target proteins. X-ray cocrystallization, hydrogen-deuterium (H / D) exchange, mass spectrometry combined with enzymatic digestion, and Several other methods known to those skilled in the art are required.

[0213] Mapping of canine IL-4 receptor alpha epitopes using mass spectrometry. : To identify epitopes recognized by anti-canine IL-4 receptor alpha mAbs, [CovalX Instrument] uses a method based on chemical crosslinking and mass spectrometry detection. [nt Incorporated]. Epitope mapping of the canine IL-4 receptor alpha chain. The application of this technology to the epitonitis recognized by the mAb listed in Table 4 is This led to the identification of the p.

[0214] Table 4 shows six antibodies used to target the canine IL-4 receptor alpha epitope. The results from the ping suggest that specific substances are present within the extracellular domain of the canine IL-4 receptor alpha. This indicates that the mAb recognizes the target peptide epitope. Notably, the test Each of the six monoclonal antibodies (mAbs) tested showed two to three specific epitopes. It was determined. Interestingly, one of the epitopes identified for mAb 2E2 is mA b. It has the exact same amino acid sequence as 11B6 (i.e., sequence number 158). This was discovered. As shown in Table 4 below, mAb:4D8, 11H2 and 11B All 6s recognize the epitope labeled "1", which is the same linear amino acid sequence. mAb:11H2, 4H3, and 2E2 are all parts of another linear amino acid sequence. Recognizing the epitope labeled "2", mAb:4H3 and 2H2 are all the same. It recognizes the epitope labeled "3," which is the linear amino acid sequence portion of 3. Related E This relative match in pitope identification is because these six monoclonal antibodies are found in dogs. The recognition of a limited number of parts within the extracellular domain of the IL-4 receptor alpha It is showing. [Table 4]

[0215] Regarding the six antibodies listed in Table 4 above, in addition to the CDR described in Example 5, Considering this, there is a one-to-one correspondence between each set of CDRs and their corresponding epitopes in Table 4. The relationship is defined. This relationship is shown in Example 5 for each of the six antibodies in Table 4. The relationship between the set of six CDRs and the corresponding epitopes to which they are linked is determined. This makes it possible. Therefore, the example described in Example 5, which binds to the corresponding epitope in Table 4, An antibody having a specific set of 6 CDRs (for example, a canine antibody) is also part of the present invention. That is the case.

[0216] Example 7 Construction of a canine-modified anti-canine IL-4 receptor alpha monoclonal antibody To perform the canine transformation operation, the DNA sequences encoding the heavy and light chains of canine IgG are used. The DNA and protein sequences of the canine heavy and light chains were determined to be publicly known in the art. This can be obtained by searching the NCBI gene and protein database. Regarding canine antibodies, there are four known IgG subtypes, namely IgG-A, IgG -B, IgG-C and IgG-D, as well as two types of light chains, namely, kappa There is a lambda. It is not limited to any particular approach, but dogs Canine compounds can be mixed in various combinations to obtain anti-canine IL-4 receptor alpha mAbs. The overall method for producing heavy and light chains includes the following scheme.

[0217] i) VH and VL domains containing the CDR of the desired anti-IL-4 receptor alpha mAb Identify the DNA sequence.

[0218] ii) Identify the H and L chain CDRs of the desired anti-IL-4 receptor mAb.

[0219] iii) Identify appropriate sequences for the H and L chains of canine IgG.

[0220] iv) DNA sequences encoding the endogenous CDR of the canine IgG H and L chains of the above sequence. Identify.

[0221] v) DNA sequences encoding endogenous canine H and L chain CDRs are used to target the desired anti-IL-4 receptor. The DNA sequence encoding the alpha CDR is replaced. Also, if desired, several i Nuframework residues are selected from the desired anti-IL-4 receptor mAb framework region. Substitution with the specified residue is also permitted.

[0222] vi) Synthesize the DNA from step (v) and clonate it into a suitable expression plasmid. The plasmid containing the desired canine H and L chains is then transfused into HEK293 cells. To sfect.

[0223] vii) Purify the expressed canine antibody from the HEK293 supernatant.

[0224] viii) Regarding the binding of purified canine antibodies to the canine IL-4 receptor alpha chain Test it.

[0225] The application of the process broadly described above is shown in Table 5 below, where the sequence numbers are listed. A set of canine H and L chain sequences was given. [Table 5]

[0226] The present invention is formed by various combinations of canine heavy chains and light chains listed in Table 5. We provide canine antibodies that are particularly strong against the canine IL-4 receptor alpha. It exhibits a bond. In certain embodiments, the heavy chain contains the amino acid sequence of SEQ ID NO: 164. The light chain contains the amino acid sequence of SEQ ID NO: 170. More detailed embodiments of this type In this case, the heavy chain is encoded by the nucleotide sequence of sequence number 163, and the light chain is sequence It is encoded by the nucleotide sequence number 169. In another embodiment, The heavy chain contains the amino acid sequence of SEQ ID NO: 166, and the light chain contains the amino acid sequence of SEQ ID NO: 172 Includes. In a more detailed embodiment of this type, the heavy chain is the nucleo of Sequence ID No. 165. The light chain is encoded by the nucleotide sequence of SEQ ID NO: 171. In yet another embodiment, the heavy chain includes the amino acid sequence of SEQ ID NO: 168. The light chain contains the amino acid sequence of SEQ ID NO: 174. More detailed embodiments of this type In this case, the heavy chain is encoded by the nucleotide sequence of sequence number 167, and the light chain is sequence Encoded by the nucleotide sequence number 173. IL-4 receptors are affected by these canine antibodies. Binding studies to alpha are described in section 8 below and are shown in Figure 4.

[0227] As described above, in order to remove the ADCC and CDC effector functions, the canine anti The Fc region of the body is based on the modified sequence of canine IgG-B. The Fc region of these antibodies is different It can be replaced with modified Fc from the canine IgG isotype, and / or These can be combined with substitute hinge regions, as described above, and 201 U.S. Provisional Patent Application No. 62 / 030,812, filed on 30 July 2014, September 3, 2014 U.S. Provisional Patent Application No. 62 / 057,541, filed on 0, dated December 16, 2014. The application is for U.S. Provisional Patent Application No. 62 / 092,496, filed on June 8, 2015. Patent application No. 62 / 172,511 and WO 2015 / 091910 (all of them) The entire contents of are incorporated herein by reference and disclosed as examples and disclosures. [ka] TIFF0007854420000029.tif214151TIFF0007854420000030.tif213152TIFF0007854420000031.tif43152

[0228] Example 8 Reactivity of canine antibodies against canine IL-4 receptor alpha The canine antibody was tested for its reactivity to the canine IL-4 receptor alpha as follows: I tried it.

[0229] 1. Coat 200 ng / well of IL-4 receptor alpha onto an immunoplate. Then, incubate the plate at 4°C overnight.

[0230] 2. The plate is placed in a phosphate buffered saline (P) containing 0.05% Tween 20. Wash three times with BS (PBST).

[0231] 3. Dilute the plate with 0.5% bovine serum albumin (BSA) in PBS at room temperature for 45 minutes. Block for up to 60 minutes.

[0232] 4. Wash the plate three times with PBST.

[0233] 5. Dilute the canine antibody in each column or row of the dilution plate from 0.3 μg / mL to 3x. Dilute it.

[0234] 6. Transfer the diluted canine antibody to each column or row of the immunoplate, and leave the plate at room temperature. Incubate for 45-60 minutes.

[0235] 7. Wash the plate three times with PBST.

[0236] 8. Dilute hose radish peroxider 1:4000 in each well of the plate. Add the ze-labeled anti-canine IgG Fc and incubate the plate at room temperature for 45-60 minutes. ru.

[0237] 9. Wash the plate three times with PBST.

[0238] 10. Place 3,3',5,5'-tetramethylbenzidine (T) into each well of the plate. Add the MB substrate and incubate the plate at room temperature for 10-15 minutes to allow color development.

[0239] 11. Add 100 μL of 1.5 M phosphoric acid to each well to stop the reaction. 54 The plate is read at 450 nm using a reference wavelength of 0 nm.

[0240] As shown in Figure 4, the binding of the following five antibodies to the IL-4 receptor alpha was investigated: 4H3 MC, c4H3 H1-L1, c4H3 H2-L2, c4H3 H3-L3, and 2G9 MC. 2G9 MC was used as a negative control antibody. 4H3 MC is a chimeric antibody consisting of the mouse variable heavy chain region and canine constant region of the 4H3 antibody disclosed herein, as well as the light chain from the mouse 4H3 antibody. c4H3 H1-L1, c4H3 H2-L2, and c4H3 H3-L3 are three canine variants of the mouse 4H3 antibody, containing specific heavy and light chains as shown in Table 5 above. 2G9 MC is a chimeric antibody consisting of the mouse variable heavy chain and canine constant region of a mouse antibody against an antigen completely unrelated to the IL-4 receptor alpha, as well as the light chain from the mouse antibody against that unrelated antigen. Consistently, 2G9 MC did not bind to the IL-4 receptor alpha, but the other four antibodies examined—4H3 MC, c4H3 H1-L1, c4H3 H2-L2, and c4H3 H3-L3—all bound relatively strongly (see Figure 4). In one embodiment, the present invention provides the following: [Item 1] Specifically for canine interleukin 4 receptor α (IL-4Rα), which contains three light chain complementarity-determining regions (CDRs), namely CDR light chain 1 (CDRL1), CDR light chain 2 (CDRL2), and CDR light chain 3 (CDRL3), and three heavy chain CDRs, namely CDR heavy chain 1 (CDRH1), CDR heavy chain 2 (CDRH2), and CDR heavy chain 3 (CDRH3). A conjugated isolated mammalian antibody or its antigen-binding fragment, wherein (a) CDRL1 contains an amino acid sequence selected from the group consisting of SEQ ID NOs: 47, 54, 48, 52, 55, 49, 50, 51, 53, 129, 130, 131; a conserved modified variant of SEQ ID NOs: 47, 54, 48, 52, 55, 49, 50, 51, 53, 129, 130, or 131; a variant of SEQ ID NOs: 47 or 54 containing the standard structural class of 1; a variant of SEQ ID NOs: 48, 52, or 55 containing the standard structural class of 2A; a variant of SEQ ID NOs: 49 containing the standard structural class of 4; a variant of SEQ ID NOs: 50, 51, or 131 containing the standard structural class of 3; and a variant of SEQ ID NOs: 53, 129, or 130 containing the standard structural class of 6; (b) CDRL2 comprises an amino acid sequence selected from the group consisting of SEQ ID NOs. 56, 57, 58, 59, 60, 61, 62, 63, 64, 132, 133, 134; a conserved modified variant of SEQ ID NOs. 56, 57, 58, 59, 60, 61, 62, 63, 64, 132, 133, or 134; and a variant of SEQ ID NOs. 56, 57, 58, 59, 60, 61, 62, 63, 64, 132, 133, or 134, wherein the variant comprises one standard structural class; (c)CDRL3 contains an amino acid sequence selected from the group consisting of SEQ ID NOs. 65, 66, 67, 68, 69, 70, 71, 72, 73, 135, 136, 137, 138, 139; conserved modified variants of SEQ ID NOs. 65, 66, 67, 68, 69, 70, 71, 72, 73, 135, 136, 137, 138, or 139; variants of SEQ ID NOs. 65, 66, 67, 68, 69, 70, 71, 72, 73, 135, 136, 137, 138, or 139 containing one standard structural class; and variants of SEQ ID NOs. 138 containing three standard structural classes; (d) CDRH1 comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 74, 75, 76, 77, 78, 79, 80, 81, 82, 140, 141, 142, 143; a conserved modified variant of SEQ ID NOs: 74, 75, 76, 77, 78, 79, 80, 81, 82, 140, 141, 142, or 143; and a variant of SEQ ID NOs: 74, 75, 76, 77, 78, 79, 80, 81, 82, 140, 141, 142, or 143, wherein the variant comprises one standard structural class; (e) CDRH2 contains an amino acid sequence selected from the group consisting of SEQ ID NOs: 83, 84, 91, 85, 88, 89, 87, 90, 144, 145, 146, 147, 148; conserved modified variants of SEQ ID NOs: 83, 84, 91, 85, 88, 89, 87, 90, 144, 145, 146, 147, 148; variants of SEQ ID NOs: 83, 144, 146, 147, or 148 containing the standard structural class of 3A; variants of SEQ ID NOs: 84, 91, or 145 containing the standard structural class of 2A; variants of SEQ ID NOs: 85, 88, or 89 containing the standard structural class of 2B; and variants of SEQ ID NOs: 87 or 90 containing the standard structural class of 1; (f) CDRH3 is a variant of SEQ ID NO: 92; a conserved modified variant of SEQ ID NO: 92; a variant of SEQ ID NO: 92 containing 12 standard structural classes; SEQ ID NO: 93; a conserved modified variant of SEQ ID NO: 93; a variant of SEQ ID NO: 93 containing 7 standard structural classes; SEQ ID NO: 94, SEQ ID NO: 96, SEQ ID NO: 149, SEQ ID NO: 151; a conserved modified variant of SEQ ID NO: 94, SEQ ID NO: 96, SEQ ID NO: 149 or SEQ ID NO: 151; a variant of SEQ ID NO: 94, SEQ ID NO: 96, SEQ ID NO: 149 or SEQ ID NO: 151 containing 15 standard structural classes; SEQ ID NO: 95; a conserved modified variant of SEQ ID NO: 95; or a variant of SEQ ID NO: 95 containing 11 standard structural classes; SEQ ID NO: 97, SEQ ID NO: 100; SEQ ID NO: 97 The amino acid sequence includes an amino acid sequence selected from the group consisting of: a conserved modified variant of SEQ ID NO: 100; a variant of SEQ ID NO: 97 or SEQ ID NO: 100 containing 6 standard structural classes; SEQ ID NO: 98; a conserved modified variant of SEQ ID NO: 98; a variant of SEQ ID NO: 98 containing 4 standard structural classes; SEQ ID NO: 99, SEQ ID NO: 153; a conserved modified variant of SEQ ID NO: 99 or SEQ ID NO: 153; a variant of SEQ ID NO: 99 or SEQ ID NO: 153 containing 13 standard structural classes; SEQ ID NO: 150; a conserved modified variant of SEQ ID NO: 150; a variant of SEQ ID NO: 150 containing 10 standard structural classes; SEQ ID NO: 152; and a conserved modified variant of SEQ ID NO: 152; and a variant of SEQ ID NO: 152 containing 9 standard structural classes. Here, the antibody and its antigen-binding fragment are canine IL-4R α An isolated mammalian antibody or its antigen-binding fragment that binds to [a specific target]. [Item 2] (a) CDRL1 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 54, SEQ ID NO: 129, SEQ ID NO: 130, SEQ ID NO: 131; Conserved modified variants of SEQ ID NO: 54, SEQ ID NO: 129, SEQ ID NO: 130 or SEQ ID NO: 131; Variants of SEQ ID NO: 54 containing 1 standard structural class; Variants of SEQ ID NO: 131 containing 3 standard structural classes; and Variants of SEQ ID NO: 129 or SEQ ID NO: 130 containing 6 standard structural classes; (b) CDRL2 comprises an amino acid sequence selected from the group consisting of SEQ ID NOs. 60, 63, 132, 133, 134; conserved modified variants of SEQ ID NOs. 60, 63, 132, 133, or 134; and variants of SEQ ID NOs. 60, 63, 132, 133, or 134, wherein the variant comprises one standard structural class; (c) CDRL3 contains an amino acid sequence selected from the group consisting of: SEQ ID NO: 72, SEQ ID NO: 135, SEQ ID NO: 136, SEQ ID NO: 137, SEQ ID NO: 138, SEQ ID NO: 139; Conserved modified variants of SEQ ID NO: 72, SEQ ID NO: 135, SEQ ID NO: 136, SEQ ID NO: 137, SEQ ID NO: 138 or SEQ ID NO: 139; Variants of SEQ ID NO: 72, SEQ ID NO: 135, SEQ ID NO: 136, SEQ ID NO: 137 or SEQ ID NO: 139 containing 1 standard structural class; and Variants of SEQ ID NO: 138 containing 3 standard structural classes; (d) CDRH1 comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 81, 140, 141, 142, 143; conserved modified variants of SEQ ID NOs: 81, 140, 141, 142, or 143; and variants of SEQ ID NOs: 81, 140, 141, 142, or 143, wherein the variant comprises one standard structural class; (e) CDRH2 contains an amino acid sequence selected from the group consisting of: SEQ ID NOs: 90, 144, 145, 146, 147, 148; conserved modified variants of SEQ ID NOs: 90, 144, 145, 146, 147, 148; variants of SEQ ID NOs: 144, 146, 147, or 148 containing the standard structural class of 3A; variants of SEQ ID NOs: 145 containing the standard structural class of 2A; and variants of SEQ ID NOs: 90 containing the standard structural class of 1; (f) CDRH3 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 149, SEQ ID NO: 151; Conserved modified variants of SEQ ID NO: 149 or SEQ ID NO: 151; Variants of SEQ ID NO: 149 or SEQ ID NO: 151 containing 15 standard structural classes; SEQ ID NO: 99, SEQ ID NO: 153; Conserved modified variants of SEQ ID NO: 99 or SEQ ID NO: 153; Variants of SEQ ID NO: 99 or SEQ ID NO: 153 containing 13 standard structural classes; SEQ ID NO: 150; Conserved modified variants of SEQ ID NO: 150; Variants of SEQ ID NO: 150 containing 10 standard structural classes; SEQ ID NO: 152; Conserved modified variants of SEQ ID NO: 152; and Variants of SEQ ID NO: 152 containing 9 standard structural classes; Here, the antibody and its antigen-binding fragment are canine IL-4R α Binds to canine interleukin 4 and canine IL-4R α An isolated mammalian antibody or antigen-binding fragment as described in item 1, which blocks the binding of the antigen. [Item 3] (a) CDRL1 comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 129, a conserved modified variant of SEQ ID NO: 129, and a variant of SEQ ID NO: 129 containing six standard structural classes; (b) CDRL2 comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 132, a conserved modified variant of SEQ ID NO: 132, and a variant of SEQ ID NO: 132 containing a standard structural class; (c) CDRL3 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 135, a conserved modified variant of SEQ ID NO: 135, and a variant of SEQ ID NO: 135 containing one standard structural class; (d) CDRH1 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 140, a conserved modified variant of SEQ ID NO: 140, and a variant of SEQ ID NO: 140 that includes a standard structural class; (e) CDRH2 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 144, a conserved modified variant of SEQ ID NO: 144, and a variant of SEQ ID NO: 144 that includes the standard structural class 3A; (f) CDRH3 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 149, a conserved modified variant of SEQ ID NO: 149, and variants of SEQ ID NO: 149 that include 15 standard structural classes; Here, the antibody and its antigen-binding fragment are canine IL-4R α Binds to canine interleukin 4 and canine IL-4R α An isolated mammalian antibody or antigen-binding fragment as described in item 2, which blocks the binding of the antigen. [Item 4] (a) CDRL1 comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 130, a conserved modified variant of SEQ ID NO: 130, and a variant of SEQ ID NO: 130 containing six standard structural classes; (b) CDRL2 comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 133, a conserved modified variant of SEQ ID NO: 133, and a variant of SEQ ID NO: 133 containing a standard structural class; (c) CDRL3 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 136, a conserved modified variant of SEQ ID NO: 136, and a variant of SEQ ID NO: 136 containing one standard structural class; (d) CDRH1 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 141, a conserved modified variant of SEQ ID NO: 141, and a variant of SEQ ID NO: 141 that includes a standard structural class; (e) CDRH2 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 145, a conserved modified variant of SEQ ID NO: 145, and a variant of SEQ ID NO: 145 that includes the standard structural class 2A; (f) CDRH3 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 150, a conserved modified variant of SEQ ID NO: 150, and variants of SEQ ID NO: 150 that include 10 standard structural classes; Here, the antibody and its antigen-binding fragment are canine IL-4R α Binds to canine interleukin 4 and canine IL-4R α An isolated mammalian antibody or antigen-binding fragment as described in item 2, which blocks the binding of the antigen. [Item 5] (a) CDRL1 comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 129, a conserved modified variant of SEQ ID NO: 129, and a variant of SEQ ID NO: 129 containing six standard structural classes; (b) CDRL2 comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 134, a conserved modified variant of SEQ ID NO: 134, and a variant of SEQ ID NO: 134 containing a standard structural class; (c) CDRL3 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 137, a conserved modified variant of SEQ ID NO: 137, and a variant of SEQ ID NO: 137 that includes one standard structural class; (d) CDRH1 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 140, a conserved modified variant of SEQ ID NO: 140, and a variant of SEQ ID NO: 140 that includes a standard structural class; (e) CDRH2 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 146, a conserved modified variant of SEQ ID NO: 146, and a variant of SEQ ID NO: 146 that includes the standard structural class 3A; (f) CDRH3 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 151, a conserved modified variant of SEQ ID NO: 151, and variants of SEQ ID NO: 151 that include 15 standard structural classes; Here, the antibody and its antigen-binding fragment are canine IL-4R α Binds to canine interleukin 4 and canine IL-4R α An isolated mammalian antibody or antigen-binding fragment as described in item 2, which blocks the binding of the antigen. [Item 6] (a) CDRL1 comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 131, a conserved modified variant of SEQ ID NO: 131, and variants of SEQ ID NO: 131 including three standard structural classes; (b) CDRL2 comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 60, a conserved modified variant of SEQ ID NO: 60, and a variant of SEQ ID NO: 60 containing a standard structural class of 1; (c) CDRL3 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 138, a conserved modified variant of SEQ ID NO: 138, and variants of SEQ ID NO: 138 that include the standard structural class of 3; (d) CDRH1 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 142, a conserved modified variant of SEQ ID NO: 142, and a variant of SEQ ID NO: 142 that includes a standard structural class; (e) CDRH2 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 147, a conserved modified variant of SEQ ID NO: 147, and a variant of SEQ ID NO: 147 that includes the standard structural class 3A; (f) CDRH3 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 152, a conserved modified variant of SEQ ID NO: 152, and a variant of SEQ ID NO: 152 that includes 9 standard structural classes; Here, the antibody and its antigen-binding fragment are canine IL-4R α Binds to canine interleukin 4 and canine IL-4R α An isolated mammalian antibody or antigen-binding fragment as described in item 2, which blocks the binding of the antigen. [Item 7] (a) CDRL1 comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 129, a conserved modified variant of SEQ ID NO: 129, and a variant of SEQ ID NO: 129 containing six standard structural classes; (b) CDRL2 comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 132, a conserved modified variant of SEQ ID NO: 132, and a variant of SEQ ID NO: 132 containing a standard structural class; (c) CDRL3 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 139, a conserved modified variant of SEQ ID NO: 139, and a variant of SEQ ID NO: 139 containing a standard structural class; (d) CDRH1 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 143, a conserved modified variant of SEQ ID NO: 143, and a variant of SEQ ID NO: 143 that includes a standard structural class; (e) CDRH2 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 148, a conserved modified variant of SEQ ID NO: 148, and a variant of SEQ ID NO: 148 that includes the standard structural class 3A; (f) CDRH3 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 153, a conserved modified variant of SEQ ID NO: 153, and a variant of SEQ ID NO: 153 that includes 13 standard structural classes; Here, the antibody and its antigen-binding fragment are canine IL-4R α Combined with dogs Canine IL-4R to Interleukin-4 α The isolated substance described in item 2 blocks the binding of Mammalian antibodies or antigen-binding fragments. [Item 8] (a) CDRL1 comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 54, a conserved modified variant of SEQ ID NO: 54, and a variant of SEQ ID NO: 54 that includes a standard structural class of 1; (b) CDRL2 comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 63, a conserved modified variant of SEQ ID NO: 63, and a variant of SEQ ID NO: 63 that includes a standard structural class of 1; (c) CDRL3 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 72, a conserved modified variant of SEQ ID NO: 72, and a variant of SEQ ID NO: 72 that includes a standard structural class of 1; (d) CDRH1 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 81, a conserved modified variant of SEQ ID NO: 81, and a variant of SEQ ID NO: 81 that includes a standard structural class of 1; (e) CDRH2 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 90, a conserved modified variant of SEQ ID NO: 90, and a variant of SEQ ID NO: 90 containing a standard structural class of 1; (f) CDRH3 contains an amino acid sequence selected from the group consisting of SEQ ID NO: 99, a conserved modified variant of SEQ ID NO: 99, and variants of SEQ ID NO: 99 that include 13 standard structural classes; Here, the antibody and its antigen-binding fragment are canine IL-4R α Combined with dogs Canine IL-4R to Interleukin-4 α The isolated substance described in item 2 blocks the binding of Mammalian antibodies or antigen-binding fragments. [Item 9] An isolated mammalian antibody or its antigen-binding fragment as described in item 1, 2, 3, 4, 5, 6, 7, or 8, wherein the mammalian antibody is a mouse antibody. [Item 10] An isolated mammalian antibody or its antigen-binding fragment, as described in item 1, 2, 3, 4, 5, 6, 7, or 8, which is a canine antibody or its canine antigen-binding fragment. [Item 11] A canine antibody or canine antigen-binding fragment according to item 10, comprising a hinge region containing an amino acid sequence selected from the group consisting of SEQ ID NO: 101, SEQ ID NO: 102, SEQ ID NO: 103, and SEQ ID NO: 104. [Item 12] An isolated mammalian antibody or its antigen-binding fragment, as described in item 6, which is a canine antibody or its canine antigen-binding fragment. [Item 13] A canine antibody or its antigen-binding fragment according to item 12, comprising a heavy chain containing an amino acid sequence selected from the group consisting of SEQ ID NO: 164, SEQ ID NO: 166, and SEQ ID NO: 168, a light chain containing an amino acid sequence selected from the group consisting of SEQ ID NO: 170, SEQ ID NO: 172, and SEQ ID NO: 174, or a combination of the heavy chain and the light chain. [Item 14] A canine antibody or its antigen-binding fragment according to item 13, comprising a heavy chain encoded by a nucleotide sequence selected from the group consisting of SEQ ID NO: 163, SEQ ID NO: 165, and SEQ ID NO: 167, a light chain encoded by a nucleotide sequence selected from the group consisting of SEQ ID NO: 169, SEQ ID NO: 171, and SEQ ID NO: 173, or a combination of the heavy chain and the light chain. [Item 15] Canine interleukin-4 receptor α (IL-4R) α Isolated milk that specifically binds to ) Antibodies or their antigen-binding fragments, including canine IL-4R α When joining, The antibody binds to at least one amino acid residue in the amino acid sequence of SEQ ID NO: 125, SEQ ID NO: 126, SEQ ID NO: 127, SEQ ID NO: 128, SEQ ID NO: 154, SEQ ID NO: 155, SEQ ID NO: 156, SEQ ID NO: 157, SEQ ID NO: 158, SEQ ID NO: 159, SEQ ID NO: 160, SEQ ID NO: 161, SEQ ID NO: 162, or any combination thereof, wherein the antibody or its antigen-binding fragment binds to canine IL-4Rα and to canine interleukin 4 NuIL-4R α Isolated mammalian antibodies or their antigen-binding fragments that block the binding nt. [Item 16] Inu IL-4R α When binding, the mammalian antibody or its antigen-binding fragment The claim is that the canine antibody or its antigen-binding fragment binds to at least one amino acid residue in the amino acid sequence of SEQ ID NO: 125, SEQ ID NO: 126, SEQ ID NO: 127, SEQ ID NO: 128, SEQ ID NO: 154, SEQ ID NO: 155, SEQ ID NO: 156, SEQ ID NO: 157, SEQ ID NO: 158, SEQ ID NO: 159, SEQ ID NO: 160, SEQ ID NO: 161, SEQ ID NO: 162 or any combination thereof, wherein the antibody or its antigen-binding fragment binds to canine IL-4Rα and blocks the binding of canine IL-4Rα to canine interleukin 4. An isolated mammalian antibody or its antigen-binding fragment as described in item 1, 2, 3, 4, 5, 6, 7, or 8, or a canine antibody or its antigen-binding fragment as described in item 10, 11, 12, 13, or 14. [Item 17] Inu IL-4R α When binding, the mammalian antibody or its antigen-binding fragment, or the canine antibody or its antigen-binding fragment, binds to at least one amino acid residue in an amino acid sequence selected from the group consisting of SEQ ID NO: 154, SEQ ID NO: 155, SEQ ID NO: 156, SEQ ID NO: 159, SEQ ID NO: 160, SEQ ID NO: 161, and any combination thereof, where the antibody or its antigen-binding fragment binds to canine IL-4Rα and to canine interleukin 4. α An isolated mammalian antibody or its antigen-binding fragment as described in item 6 or 7, or a canine antibody or its antigen-binding fragment as described in item 12, 13 or 14, which blocks the binding of the antibody. [Item 18] The mammalian antibody or its antigen-binding fragment, or the canine antibody or its antigen-binding fragment, has the following characteristics: (i)1×10 -5 M~1×10 -12 Canine IL-4R with M's dissociation constant (Kd) α To join; (ii) 1 × 10 2 M -1 s -1 ~1×10 7 M -1 s -1 On speed (k on ) with the IL-4R α To join; (iii) 1 × 10 -3 s -1 ~1×10 -8 s-1 Off speed (k off ) with the IL-4R α To join; (iv) Blocking the binding of IL-4 to type I IL-4 receptors; (v) Blocking the binding of IL-13 to the type I IL-4 receptor; (vi) Blocking the binding of IL-4 to the type II IL-4 receptor; and (vii) Blocking the binding of IL-13 to type II IL-4 receptors. A mammalian antibody or its antigen-binding fragment as described in item 1, 2, 3, 4, 5, 6, 7, 8, 15, 16, or 17, representing one, two, three, four, five, six, or all of the above, or a canine antibody or its antigen-binding fragment as described in item 10, 11, 12, 13, 14, 16, or 17. [Item 19] Inu IL-4R α Regarding binding to, one or more mammalian antibodies or their antigen-binding fragments described in item 1, 2, 3, 4, 5, 6, 7, 8, 15, 16, 17, or 18, or a canine monoclonal antibody or its antigen-binding fragment described in item 10, 11, 12, 13, 14, 16, 17, or 18, which cross-compete with canine IL-4Rα and canine IL-4Rα α A canine monoclonal antibody or its antigen-binding fragment that blocks the binding of the antigen. [Item 20] A mammalian antibody or its antigen-binding fragment as described in item 1, 2, 3, 4, 5, 6, 7, 8, 15, 16, 17, or 18, a mouse antibody or its antigen-binding fragment as described in item 9, or an isolated nucleic acid encoding the light chain of a canine antibody or its antigen-binding fragment as described in item 10, 11, 12, 13, 14, 16, 17, 18, or 19. [Item 21] A mammalian antibody or its antigen-binding fragment as described in item 1, 2, 3, 4, 5, 6, 7, 8, 15, 16, 17, or 18, a mouse antibody or its antigen-binding fragment as described in item 9, or an isolated nucleic acid encoding the heavy chain of a canine antibody or its antigen-binding fragment as described in item 10, 11, 12, 13, 14, 16, 17, 18, or 19. [Item 22] Sequence numbers 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 1 Isolated nucleic acids as described in item 20 or 21, encoding one or more amino acid sequences selected from the group consisting of 29, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, and 153. [Item 23] An expression vector containing an isolated nucleic acid as described in item 20, 21, or 22. [Item 24] Host cells containing the expression vector described in item 23. [Item 25] An isolated peptide containing 5 to 25 amino acid residues that is 95% identical to an amino acid sequence selected from the group consisting of SEQ ID NOs: 125, 126, 127, 128, 154, 155, 156, 157, 158, 159, 160, 161, and 162, and that binds to an isolated mammalian antibody or its binding fragment described in item 1, 2, 3, 4, 5, 6, 7, 8, 15, 16, 17, or 18, or to a canine antibody or its antigen-binding fragment described in item 10, 11, 12, 13, 14, 16, 17, 18, or 19. [Item 26] A fusion protein containing the peptide described in item 25. [Item 27] A fusion protein as described in item 26, further comprising the Fc region of non-canine mammalian IgG. [Item 28] Isolated nucleic acids encoding an isolated peptide as described in item 25, a fusion protein as described in item 26, or a fusion protein as described in item 27. [Item 29] An expression vector containing the isolated nucleic acid described in item 28. [Item 30] Host cells containing the expression vector described in item 29. [Item 31] A pharmaceutical composition comprising a canine antibody as described in item 10, 11, 12, 13, 14, 16, 17, 18, or 19, or a nucleic acid as described in item 20, 21, 22, or 28, an expression vector as described in item 23 or 29, an isolated peptide as described in item 25, a fusion protein as described in item 26 or 27, or any combination thereof, and a pharmaceutically acceptable carrier or diluent. [Item 32] A method for reducing the activity of immune cells, comprising administering a therapeutically effective amount of the pharmaceutical composition described in item 31 to a subject requiring a reduction in the activity of immune cells. [Item 33] (i) Treatment of atopic dermatitis, or (ii) Treatment of asthma, or (iii) Treatment of atopic dermatitis and treatment of asthma The method used, as described in item 32.

Claims

1. Inu IL-4R α An isolated mammalian antibody or its antigen-binding fragment that specifically binds to, The antibody comprises three light chain complementarity-determining regions (CDRs), namely CDR light chain 1 (CDRL1), CDR light chain 2 (CDRL2), and CDR light chain 3 (CDRL3), and three heavy chain CDRs, namely CDR heavy chain 1 (CDRH1), CDR heavy chain 2 (CDRH2), and CDR heavy chain 3 (CDRH3); (a) CDRL1 contains the amino acid sequence of SEQ ID NO: 129; (b) CDRL2 contains the amino acid sequence of SEQ ID NO: 132; (c) CDRL3 contains the amino acid sequence of SEQ ID NO: 135; (d) CDRH1 contains the amino acid sequence of SEQ ID NO: 140; (e) CDRH2 contains the amino acid sequence of SEQ ID NO: 144; and (f) CDRH3 contains the amino acid sequence of SEQ ID NO: 149; Here, the antibody and its antigen-binding fragment are canine IL-4R α It binds to canine interleukin 4 and canine IL-4R α The isolated mammalian antibody or antigen-binding fragment that blocks the binding of the antibody.

2. Inu IL-4R α An isolated mammalian antibody or its antigen-binding fragment that specifically binds to, The antibody comprises three light chain complementarity-determining regions (CDRs), namely CDR light chain 1 (CDRL1), CDR light chain 2 (CDRL2), and CDR light chain 3 (CDRL3), and three heavy chain CDRs, namely CDR heavy chain 1 (CDRH1), CDR heavy chain 2 (CDRH2), and CDR heavy chain 3 (CDRH3); (a) CDRL1 contains the amino acid sequence of SEQ ID NO: 130; (b) CDRL2 contains the amino acid sequence of SEQ ID NO: 133; (c) CDRL3 contains the amino acid sequence of SEQ ID NO: 136; (d) CDRH1 contains the amino acid sequence of SEQ ID NO: 141; (e) CDRH2 contains the amino acid sequence of SEQ ID NO: 145; and (f) CDRH3 contains the amino acid sequence of SEQ ID NO: 150; Here, the antibody and its antigen-binding fragment are canine IL-4R α It binds to canine interleukin 4 and canine IL-4R α The isolated mammalian antibody or antigen-binding fragment that blocks the binding of the antibody.

3. Inu IL-4R α An isolated mammalian antibody or its antigen-binding fragment that specifically binds to, The antibody comprises three light chain complementarity-determining regions (CDRs), namely CDR light chain 1 (CDRL1), CDR light chain 2 (CDRL2), and CDR light chain 3 (CDRL3), and three heavy chain CDRs, namely CDR heavy chain 1 (CDRH1), CDR heavy chain 2 (CDRH2), and CDR heavy chain 3 (CDRH3); (a) CDRL1 contains the amino acid sequence of SEQ ID NO: 129; (b) CDRL2 contains the amino acid sequence of SEQ ID NO: 134; (c) CDRL3 contains the amino acid sequence of SEQ ID NO: 137; (d) CDRH1 contains the amino acid sequence of SEQ ID NO: 140; (e) CDRH2 contains the amino acid sequence of SEQ ID NO: 146; and (f) CDRH3 contains the amino acid sequence of SEQ ID NO: 151; Here, the antibody and its antigen-binding fragment bind to canine IL-4R α and block the binding of canine IL-4R α to canine interleukin 4, said isolated mammalian antibody or antigen-binding fragment.

4. Inu IL-4R α An isolated mammalian antibody or its antigen-binding fragment that specifically binds to, The antibody comprises three light chain complementarity-determining regions (CDRs), namely CDR light chain 1 (CDRL1), CDR light chain 2 (CDRL2), and CDR light chain 3 (CDRL3), and three heavy chain CDRs, namely CDR heavy chain 1 (CDRH1), CDR heavy chain 2 (CDRH2), and CDR heavy chain 3 (CDRH3); (a) CDRL1 contains the amino acid sequence of SEQ ID NO: 129; (b) CDRL2 contains the amino acid sequence of SEQ ID NO: 132; (c) CDRL3 contains the amino acid sequence of SEQ ID NO: 139; (d) CDRH1 contains the amino acid sequence of SEQ ID NO: 143; (e) CDRH2 contains the amino acid sequence of SEQ ID NO: 148; and (f) CDRH3 contains the amino acid sequence of SEQ ID NO: 153; Here, the antibody and its antigen-binding fragment are canine IL-4R α It binds to canine interleukin 4 and canine IL-4R α The isolated mammalian antibody or antigen-binding fragment that blocks the binding of the antibody.

5. Inu IL-4R α An isolated mammalian antibody or its antigen-binding fragment that specifically binds to, The antibody comprises three light chain complementarity-determining regions (CDRs), namely CDR light chain 1 (CDRL1), CDR light chain 2 (CDRL2), and CDR light chain 3 (CDRL3), and three heavy chain CDRs, namely CDR heavy chain 1 (CDRH1), CDR heavy chain 2 (CDRH2), and CDR heavy chain 3 (CDRH3); (a) CDRL1 contains the amino acid sequence of SEQ ID NO: 54; (b) CDRL2 contains the amino acid sequence of SEQ ID NO: 63; (c) CDRL3 contains the amino acid sequence of SEQ ID NO: 72; (d) CDRH1 contains the amino acid sequence of SEQ ID NO: 81; (e) CDRH2 contains the amino acid sequence of SEQ ID NO: 90; and (f) CDRH3 contains the amino acid sequence of SEQ ID NO: 99; Here, the antibody and its antigen-binding fragment are canine IL-4R α It binds to canine interleukin 4 and canine IL-4R α The isolated mammalian antibody or antigen-binding fragment that blocks the binding of the antibody.

6. The isolated mammalian antibody or its antigen-binding fragment according to claim 1, 2, 3, 4, or 5, wherein the mammalian antibody is a mouse antibody.

7. The isolated mammalian antibody or its antigen-binding fragment according to claim 1, 2, 3, 4, or 5, wherein the mammalian antibody is a canine antibody or a canine antigen-binding fragment thereof.

8. The isolated mammalian antibody or its antigen-binding fragment according to claim 7, further comprising a hinge region containing an amino acid sequence selected from the group consisting of SEQ ID NO: 101, SEQ ID NO: 102, SEQ ID NO: 103, and SEQ ID NO:

104.

9. Inu IL-4R α When binding, the isolated mammalian antibody or its antigen-binding fragment or the canine antibody or its antigen-binding fragment binds to at least one amino acid residue in the amino acid sequence of SEQ ID NO: 125, SEQ ID NO: 126, SEQ ID NO: 127, SEQ ID NO: 128, SEQ ID NO: 154, SEQ ID NO: 155, SEQ ID NO: 156, SEQ ID NO: 157, SEQ ID NO: 158, SEQ ID NO: 159, SEQ ID NO: 160, SEQ ID NO: 161, SEQ ID NO: 162 or any combination thereof, wherein the antibody or its antigen-binding fragment binds to canine IL-4Rα and blocks the binding of canine IL-4Rα to canine interleukin 4, as described in claim 1, 2, 3, 4, 5, 6, 7, or 8.

10. The mammalian antibody or its antigen-binding fragment has the following characteristics: (i) 1 × 10 －５ M~1 x 10 －１２ In canine IL-4R with M's dissociation constant (Kd) α To join; (ii) 1 x 10 ２ M －１ s －１ ~1 x 10 ７ M －１ s －１ On speed (k ｏｎ ) with Inu IL-4R α To join; (iii) 1 x 10 －３ s －１ ~1 x 10 －８ s －１ Off speed (k ｏｆｆ ) with Inu IL-4R α To join; (iv) Blocking the binding of IL-4 to the type I IL-4 receptor; (v) Blocking the binding of IL-13 to the type I IL-4 receptor; (vi) Blocking the binding of IL-4 to the type II IL-4 receptor; and (vii) Blocking the binding of IL-13 to the type II IL-4 receptor. An isolated mammalian antibody or its antigen-binding fragment according to claim 1, 2, 3, 4, 5, 6, 7, 8, or 9, representing one, two, three, four, five, six, or all of the above.

11. Inu IL-4R α With respect to binding to, a canine monoclonal antibody or its antigen-binding fragment that cross-competes with one or more isolated mammalian antibodies or their antigen-binding fragments according to claim 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10, wherein it binds to canine IL-4Rα and to canine IL-4R α A canine monoclonal antibody or its antigen-binding fragment that blocks the binding of the antigen.

12. An isolated mammalian antibody or its antigen-binding fragment according to claim 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10, or an isolated nucleic acid encoding the light chain and heavy chain of a canine monoclonal antibody or its antigen-binding fragment according to claim 11.

13. A pair of isolated nucleic acids comprising: a first nucleic acid encoding the light chain of an isolated mammalian antibody or its antigen-binding fragment according to claim 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 or a canine antibody or its antigen-binding fragment according to claim 11; and a second nucleic acid encoding the heavy chain of an isolated mammalian antibody or its antigen-binding fragment according to claim 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 or a canine antibody or its antigen-binding fragment according to claim 11.

14. The isolated nucleic acid according to claim 12, or the pair of isolated nucleic acids according to claim 13, wherein the isolated nucleic acid, or the first or second nucleic acid of the pair of nucleic acids, encodes one or more amino acid sequences selected from the group consisting of SEQ ID NOs: 54, 60, 63, 72, 81, 90, 99, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, and 153.

15. An expression vector system comprising an isolated nucleic acid according to claim 12 or 14, or a pair of isolated nucleic acids according to claim 13 or 14.

16. A host cell comprising the expression vector system according to claim 15.

17. A pharmaceutical composition comprising an isolated mammalian antibody or its antigen-binding fragment according to claim 7 or 8, a canine monoclonal antibody or its antigen-binding fragment according to claim 11, an isolated nucleic acid according to claim 12 or 14, a pair of isolated nucleic acids according to claim 13 or 14, an expression vector system according to claim 15, or any combination thereof, and a pharmaceutically acceptable carrier or diluent.

18. The pharmaceutical composition according to claim 17 for use in the treatment of atopic dermatitis and / or asthma.

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