CN110300763A - For treating the anti-human CXCR3 antibody of leucoderma - Google Patents

Abstract

Provide the method and purposes of anti-CXCR3 Antybody therapy CXCR3 associated disorders such as leucoderma.In certain embodiments, the anti-CXCR3 antibody is Humanized anti-human CXCR3 antibody, has the effector function of enhancing for the cell for expressing CXCR3 on the surface.

Description

Anti-human CXCR3 antibody for the treatment of vitiligo

Cross References to Related Applications

This application claims the benefit of U.S. Provisional Patent Application No. 62/437,889, filed December 22, 2016, and European Patent Application No. 17306770.3, filed December 14, 2017, the entire contents of which are incorporated herein by reference.

Background technique

The present invention relates to antibodies and methods of using the antibodies to treat disorders associated with CXCR3 signaling, such as vitiligo.

Vitiligo is a disfiguring autoimmune skin disease in which autoreactive CD8 ⁺ T cells kill melanocytes in the epidermis, resulting in patchy hyperpigmentation. Although it is one of the most common autoimmune diseases, affecting 1 percent of the world's population, there are currently no FDA-approved treatments for vitiligo.

C-X-C motif chemokine receptor 3 (CXCR3) is a chemokine receptor mainly expressed on antigen-experiencing T cells (memory T cells), effector T cells, and activated T cells, and is involved in the transfer of these T cells to Clusters are recruited to sites of tissue inflammation in response to their major ligands: CXCL9 (MIG), CXCL10 (IP-10) and CXCL11 (I-TAC). CXCR3 and CXCL10 are expressed in human T1D patients (Uno et al, Endocr J 57:991-96 (2010); Roep et al, Clin Exp Immunol 159:338-43 (2003); Tanaka et al, Diabetes 58:2285-2291 (2009 )). In these patients, CXCL10 was expressed in the remaining insulin-producing beta cells in the islets. CXCR3 is expressed on invading T cells surrounding the islets. Similar expression patterns have been reproduced in non-obese diabetic (NOD) mice, a mouse model of type 1 diabetes (Morimoto et al., J Immunol 173:7017-24 (2004); Li et al., World J Gastroenterol 11(30) :4750-2 (2005); Sarkar et al., Diabetes 61(2):436-46 (2012)).

CXCR3 is expressed by cutaneous CD3 ⁺ lymphocytes and plasmacytoid dendritic cells, and its chemokine ligands CXCL10 and CXCL9 are upregulated in psoriatic lesions (Rottman et al., Lab Invest 81(3):335-47 (2001) ; Chen et al., Arch Dermatol Res 302(2):113-23(2010)). CXCR3 is also expressed on infiltrating T cells present in certain types of inflamed tissues, whereas CXCL9, CXCL10 and CXCL11 are often produced by localized cells in inflammatory lesions.

Upregulation of CXCR3 has been implicated in a range of autoimmune disorders. CXCR3 expression is largely absent in naive T cells and is upregulated after activation with antigen. CXCR3 recruits these cells, including T helper type 1 (Th1 ), to sites of tissue inflammation in response to its primary ligand. β cells in Langerhans islets express CXCL9 and CXCL10 (Frigerio et al., Nat Med 8:1414-20 (2002)), whereas T cells that have infiltrated the pancreas express CXCR3 (Christen et al., J Immunol 171:6838-45 (2003 ); Van Halteren et al, Diabetologia 48:75-82 (2005); Uno et al, Endocr J 57:991-96 (2010); Roep et al, Clin Exp Immunol 159:338-43 (2003); Tanaka et al, Diabetes 58 :2285-2291 (2009); Sarkar et al., Diabetes 61(2):436-46 (2012)). US Patent No. 8,865,870 to Youd et al. describes anti-CXCR3 antibodies.

Given the prevalence of vitiligo and other disorders involving CXCR3, additional approaches to target CXCR3 signaling, eg, to treat or reduce the progression of disorders such as vitiligo in patients, are needed.

Contents of the invention

Provided herein are methods of treating CXCR3-related diseases and disorders, such as vitiligo, by administering a humanized antibody or antigen-binding fragment thereof that specifically binds CXCR3. In certain embodiments, the anti-CXCR3 antibodies provided for the methods and uses herein have the ability to direct the depletion of CXCR3-expressing cells, or can be engineered to have an enhanced ability to direct the depletion of CXCR3-expressing cells for the treatment of CXCR3-related diseases and Disorders (such as Vitiligo).

In some embodiments, the humanized antibodies or antigen-binding fragments thereof provided for the methods and uses herein have at least 0.885 (when comparing all residues of the VH and VL chains except the residues of the D region of VH) or at least Germinal ability score of 0.950 (when only residues of the framework regions as determined by IMTG) were compared. In some embodiments, a humanized antibody or antigen-binding fragment thereof provided herein has a KD of at least 1 x 10 ^-9 M. In some embodiments, a humanized antibody or antigen-binding fragment thereof provided herein has a kd of less than 7 x 10 ⁻⁵ 1/Ms. In some embodiments, the humanized antibodies or antigen-binding fragments thereof provided herein have a germinal ability score (when comparing all residues of the VH and VL chains except the D region residues of VH) of at least 0.885 or at least 0.950 (when comparing only residues of the framework region as determined by IMTG) and a KD of at least 1 x 10 ^-9 M, and at least 0.885 (when comparing residues of the VH and VL chains other than the D region residues of VH) all residues) or a germinability score of at least 0.950 (when comparing only residues of the framework regions as determined by IMTG). In some embodiments, the humanized antibodies or antigen-binding fragments thereof provided herein have at least 0.885 (when comparing all residues of the VH and VL chains except the D region residues of VH) or at least 0.950 (when comparing only A germinability score for framework region residues as determined by IMTG, a KD of at least 1 x 10 ^-9 M, and a kd of less than 7 x 10 ^-5 1/Ms.

In certain embodiments, humanized CXCR3 antibodies comprising a particular light chain variable region paired with a particular heavy chain variable region are provided. In certain embodiments, the humanized CXCR3 antibodies provided herein comprise a variant human IgGl Fc region that confers enhanced effector function against cells expressing human CXCR3 on their surface.

In a first aspect, provided herein is a humanized anti-human CXCR3 antibody or a pharmaceutical composition thereof for use in a method of depleting CXCR3 expressing cells in a subject, wherein the humanized anti-human CXCR3 antibody comprises A heavy chain (HC) with a heavy chain variable region (VH) and a light chain (LC) with a light chain variable region (VL). In some embodiments, CD4+ T cells are depleted. In some embodiments, CD8+ T cells are depleted. In some embodiments, CD4+ T cells and CD8+ T cells are depleted. In some embodiments, CD4+ memory T cells are depleted. In some embodiments, CD8+ memory T cells are depleted. In some embodiments, CD4+ memory T cells and CD8+ memory T cells are depleted. In one embodiment, the subject has a T cell mediated autoimmune disease. In another embodiment, the subject has vitiligo.

In one embodiment of the first aspect, the VH and VL of the humanized anti-human CXCR3 antibody provided herein comprise the amino acid sequences of the sequence pairs shown in Table 1, and the HC further comprises a human IgG1 Fc region, the human The IgG1 Fc region comprises the amino acid sequence of any one of SEQ ID NO: 2, 3, 4, 5, 6, 7, 8, 9, 10 or 11.

Table 1

In another embodiment of the first aspect, the VH of the humanized anti-human CXCR3 antibody provided herein comprises the amino acid sequence of SEQ ID NO:20, and the VL comprises the amino acid sequence of SEQ ID NO:24. In some embodiments, the humanized human IgG1 Fc region comprises the amino acid sequence of SEQ ID NO:2, or SEQ ID NO:9, or SEQ ID NO:10, or SEQ ID NO:11.

In another embodiment of the first aspect, the VH of the humanized anti-human CXCR3 antibody provided herein comprises the amino acid sequence of SEQ ID NO:18, and the VL comprises the amino acid sequence of SEQ ID NO:22. In some embodiments, the antibody comprises a human IgG1 Fc region. In some embodiments, the human IgG1 Fc region comprises the amino acid sequence of SEQ ID NO:2, or SEQ ID NO:9, or SEQ ID NO:10, or SEQ ID NO:11.

In other embodiments of the first aspect, the HC and LC of the humanized anti-human CXCR3 antibodies provided herein comprise the amino acid sequences of the pairs of SEQ ID NOs shown in Table 2.

Table 2

In another embodiment of the first aspect, the HC of the humanized anti-human CXCR3 antibody provided herein comprises a human IgG1 Fc region with reduced fucose content. In some embodiments, provided humanized anti-human CXCR3 antibodies are produced in host cells cultured in medium containing a glycosylation inhibitor. In one embodiment, the glycosylation inhibitor is kifunensine.

In a second aspect, provided herein is a humanized anti-human CXCR3 antibody or a pharmaceutical composition thereof for use in a method of treating a T cell-mediated autoimmune disease, wherein the humanized anti-human CXCR3 antibody comprises A heavy chain (HC) with a heavy chain variable region (VH) and a light chain (LC) with a light chain variable region (VL). In one embodiment, the T cell mediated disease is vitiligo.

In a third aspect, provided herein is a method of treating a T cell-mediated autoimmune disease comprising combining a heavy chain (HC) having a heavy chain variable region (VH) and a light chain variable region (VL) comprising The humanized anti-human CXCR3 antibody of the light chain (LC) of the above is administered to a subject in need thereof. In some embodiments, the T cell-mediated autoimmune disease is vitiligo.

In a fourth aspect, provided herein is a method of treating a T cell-mediated autoimmune disease comprising providing a method comprising a heavy chain (HC) having a heavy chain variable region (VH) and a light chain variable region (VL Instructions for administering a humanized anti-human CXCR3 antibody of the light chain (LC) of ) to a subject in need thereof. In some embodiments, the T cell-mediated autoimmune disease is vitiligo.

In a fifth aspect, provided herein is a kit for treating a T cell-mediated autoimmune disease comprising a heavy chain (HC) having a heavy chain variable region (VH) and a light chain variable region ( A humanized anti-human CXCR3 antibody of the light chain (LC) of VL), and instructions for administering the humanized anti-human CXCR3 to a human subject in need thereof.

In some embodiments of the first, second, third, fourth and fifth aspects provided herein, the VH and VL of the humanized anti-human CXCR3 antibody comprise the amino acid sequences of the sequence pairs shown in Table 1, And the HC comprises a human IgG1 Fc region comprising the amino acid sequence of SEQ ID NO: 2, 3, 4, 5, 6, 7, 8, 9, 10 or 11. In another embodiment, the VH comprises the amino acid sequence of SEQ ID NO:20 and the VL comprises the amino acid sequence of SEQ ID NO:24. In another embodiment, VH comprises the amino acid sequence of SEQ ID NO:20 and VL comprises the amino acid sequence of SEQ ID NO:24, and the human IgG1 Fc region comprises SEQ ID NO:2, or SEQ ID NO:9, or SEQ ID NO:9, or SEQ ID NO:24 The amino acid sequence of ID NO: 10, or SEQ ID NO: 11.

In other embodiments of the second, third, fourth, fifth, sixth and seventh aspects provided herein, the VH comprises the amino acid sequence of SEQ ID NO: 18 and the VL comprises the amino acid sequence of SEQ ID NO: 22 sequence. In some embodiments, the humanized human IgG1 Fc region comprises the amino acid sequence of SEQ ID NO:2, or SEQ ID NO:9, or SEQ ID NO:10, or SEQ ID NO:11.

In other embodiments of the first, second, third, fourth and fifth aspects provided herein, the HC and LC of the antibody comprise the amino acid sequences of the sequence pairs shown in Table 2.

In other embodiments of the first, second, third, fourth and fifth aspects provided herein, the HC of the humanized anti-human CXCR3 antibody provided herein comprises a human IgG1 Fc with reduced fucose content Area. In some embodiments, provided humanized anti-human CXCR3 antibodies are produced in host cells cultured in medium containing a glycosylation inhibitor. In some embodiments, the glycosylation inhibitor is a Kiff base.

Description of drawings

Figure 1 is a series of six graphs showing the amount of each T cell subset in the blood following antibody treatment: "Hamster CXCR3-173" (hamster anti-mouse CXCR3), "CXCR3 mIgG2a Dab" (engineered to hamster CXCR3-173 mutated to replace mouse IgG2a constant region to remove effector function), "CXCR3 mIgG2a WT" (hamster CXCR3-173 engineered to replace mouse wild-type IgG2a constant region), "CXCR3 mIgG1-agly" (hamster CXCR3-173 engineered to replace the mouse IgG1 constant region with the N297G mutation), and "untreated" (representing an untreated control).

Figure 2A is a series of six graphs depicting the binding of recombinant mouse FcyRI (mFcRI) to various Fc-engineered forms of CXCR3-173 as measured by a surface plasmon resonance (Biacore) binding assay. BMP5, anti-BMP5 mIgG1 isotype control; mIgG1 agly, engineered to replace CXCR3-173 of the mouse IgG1 constant region with the N297G mutation; mIgG2aWT, engineered to replace CXCR3-173 of the mouse wild-type IgG2a constant region; mIgG2a Dab, CXCR3-173 engineered to replace the constant region of mouse IgG2a with mutations to remove effector function; mIgG3, CXCR3-173 engineered to replace the constant region of mouse wild-type IgG3; Hamster CXCR3, parental hamster mAb CXCR3 -173.

Figure 2B is a series of six graphs depicting the binding of recombinant mouse FcγRIIb (mFcRIIb) to various Fc-engineered forms of CXCR3-173 as measured by Biacore binding assay. The names of the antibodies are the same as in Fig. 2A.

Figure 2C is a series of six graphs depicting the binding of recombinant mouse FcγRIII (mFcRIII) to various Fc-engineered forms of CXCR3-173 as measured by Biacore binding assay. The names of the antibodies are the same as in Fig. 2A.

Figure 2D is a series of six graphs depicting the binding of recombinant mouse FcyRIV (mFcRIV) to various Fc-engineered forms of CXCR3-173 as measured by Biacore binding assay. The names of various antibodies are the same as in Fig. 2A.

Figure 3A is a table summarizing the structure-effector function characteristics of anti-human CXCR3 antibodies with engineered human IgGl constant regions.

Figure 3B is a bar graph depicting in vitro antibody-dependent cellular cytotoxicity (ADCC) of CHO-human CXCR3 target cells with various anti-human CXCR3 antibodies at the indicated concentrations and a 5:1 effector ratio to target (E:T). )-mediated cleavage. Effector cells are natural killer (NK) cells from a single donor. IgG, human IgGl isotype control. Anti-human CXCR3 mAbs tested were clone 4 (CXCR3 CL4), clone 12 (CXCR3 CL12), clone 82 (CXCR3 CL82), clone 135 (CXCR3 CL135), 53Hu37, and the engineered Fc variant of 53Hu37 as described in Figure 3A Body M1, M2 and M3. Kif, Kif base treatment. ALEM, alemtuzumab.

Figure 3C is a bar graph depicting in vitro ADCC-mediated lysis of CHO-human CXCR3 target cells with various antibodies at the indicated concentrations and 3:1 effector to target (E:T). Effector cells were from an NK-like cell line (NK92-CD16V). IgG, human IgGl isotype control. The anti-human CXCR3 mAbs tested were 53Hu37 and the engineered Fc variant M1 as shown in Figure 3A. CXCR3 CL4, anti-human CXCR3 mAb clone 4. Kif, Kif base treatment. ALEM is alemtuzumab.

Figure 4 is a table summarizing 53Hu37 and indications showing binding affinity KD for human FcγRIIa (rhFcγRIIa), human FcγRIII-F158 (rhFcγRIII-F158), human FcγRIII-V158 (rhFcγRIII-V158) and mouse FcγRIV (rmFcγRIV) Biacore data for variants. M1-M3 are as described in Figure 3A, and kif is defucosylated 53Hu37.

Figure 5A is a series of six graphs depicting the depletion of indicated T cell subsets in cynomolgus monkeys treated with indicated antibodies administered at a dose of 2 mg/kg body weight. M1: S239D/I332E variant of 53Hu37; Kif: defucosylated 53Hu37; Veh: vehicle control. CXCR3 antibody group N=8. Vehicle set N=6.

Figure 5B is a graph depicting combined pharmacokinetic data evaluating concentrations of the indicated antibodies in serum of cynomolgus monkeys previously treated with a single dose of the indicated antibodies given at a dose of 2 mg/kg body weight as indicated amount of time. Anti-human CXCR3 mAbs tested were 53Hu37, Kiff base-treated 53Hu37 (53Hu37 kif) and the M1 variant of 53Hu37 (53Hu37 M1 ).

Figure 6 is a table showing SEQ ID Nos and corresponding sequences.

Detailed ways

Reference will now be made in detail to certain exemplary embodiments in accordance with the present disclosure, some examples of which are illustrated in the accompanying drawings.

Provided herein are methods and uses of humanized antibodies or antigen-binding fragments thereof that specifically bind human CXCR3. In certain embodiments, the anti-CXCR3 antibodies provided herein have the ability to direct depletion of CXCR3 expressing cells, or are engineered to have enhanced ability to direct depletion of CXCR3 expressing cells for the treatment of CXCR3-related diseases and disorders. In some embodiments, therapies for targeting CXCR3 for the treatment of vitiligo are disclosed

Antibody

As used herein, the term "antibody" refers to an immunoglobulin molecule comprising four polypeptide chains, two heavy (H) chains and two light (L) chains interconnected by disulfide bonds, and multimers thereof ( such as IgM). Each heavy chain is comprised of a heavy chain variable region (abbreviated _VH or VH) and a heavy chain constant region ( _CH or CH). The heavy chain constant region comprises three domains, _CH1 , _CH2 and _CH3 . The Fc portion of the heavy chain comprises _CH2 and _CH3 .

Each light chain is comprised of a light chain variable region (abbreviated _VL ) and a light chain constant region ( _CL or CL). The light chain constant region comprises one domain ( _CL1 ).

The _VH and _VL regions can be further subdivided into regions of high variability, called complementarity determining regions (CDRs), interspersed with more conserved regions, called framework regions (FRs). Each _VH and _VL consists of three CDRs and four FRs, arranged from amino-terminus to carboxy-terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, and FR4.

As used herein, the term "antigen-binding fragment" of an antibody includes any naturally occurring, enzymatically obtainable, synthetic or genetically engineered polypeptide or glycoprotein that specifically binds an antigen to form a complex. Antigen-binding fragments of antibodies may be derived, for example, from intact antibody molecules using any suitable standard technique, such as proteolytic digestion or recombinant genetic engineering techniques involving manipulation and expression of DNA encoding antibody variable and optionally constant domains. Non-limiting examples of antigen binding moieties include: (i) Fab fragments; (ii) F(ab' ₎ fragments; (iii) Fd fragments; (iv) Fv fragments; (v) single chain Fv (scFv) molecules; (vi) dAb fragments; (vii) minimal recognition units (eg, isolated complementarity determining regions (CDRs)) comprising amino acid residues that mimic the hypervariable regions of antibodies. Other engineered molecules, such as bispecific antibodies, trispecific antibodies, tetraspecific antibodies and minibodies, are also included within the expression "antigen-binding fragment".

In certain embodiments, a CXCR3 antibody or antigen-binding fragment comprises at least one antigen-binding domain. In some embodiments, the antibody or fragment is multispecific, comprising two or more (e.g., 2, 3, 4, 5 or more) antigen binding domains such that the antibody or fragment is capable of binding the same or different Two or more CXCR3 molecules with epitopes, or capable of binding CXCR3 and at least one other antigen with high affinity. An antigen binding portion may comprise one or more antibody fragments that retain the ability to specifically bind an antigen. These fragments may include the heavy chain variable region and/or the light chain variable region from the parent antibody or from a variant of the parent antibody.

As used herein, the term "antigen" refers to a binding site or epitope recognized by an antibody or antigen-binding fragment thereof.

An "epitope" or "antigenic determinant" is a part of an antigenic molecule that is responsible for the specific interaction with the antigen-binding domain of an antibody.

As used herein, "binding" with respect to an antibody or antigen-binding fragment thereof means that the antibody or antigen-binding fragment thereof forms one or more non-covalent Ability to covalently bond. An antigen can be an isolated antigen, or it can exist in association with another entity, such as in the context of a polypeptide on the surface of a cell.

As used herein, the term "specifically binds" refers to an antibody or antigen ^- binding fragment thereof with a Kd of at least about 1x 10 ^-6 M, 1x 10 ^-7 M, 1x 10 ^-8 M, 1x 10 ^-9 M, 1x 10 ^{- 10} M, 1x 10 ^-11 M, 1x 10 ^-12 M or higher antigen binding capacity. In certain embodiments, the term refers to the ability of an antibody or antigen-binding fragment thereof to bind an antigen with an affinity at least 2-fold higher than its affinity for a non-specific antigen. However, it is understood that an antibody or antigen-binding fragment thereof is capable of specifically binding two or more sequence-related antigens (eg, human and cynomolgus CXCR3). Nonspecific binding typically has low affinity and moderate to high capacity. If necessary, non-specific binding can be reduced without substantially affecting specific binding by altering the binding conditions. These conditions are known in the art, and suitable conditions can be selected by one skilled in the art using conventional techniques. The conditions are usually defined in terms of antibody concentration, ionic strength of the solution, temperature, allowed binding time and concentration of blocking molecules such as serum albumin and casein.

Affinity constants can be determined by standard kinetic methods of antibody reactions, such as immunoassays (eg, ELISA) or surface plasmon resonance (SPR). Instruments and methods for detecting and monitoring association rates in real time are known and commercially available (eg, Biacore 2000, Biacore AB, Upsala, Sweden and GE Healthcare LifeSciences).

As used herein, "complementarity determining region" or "CDR" refers to one of the portions within each variable region of an antibody or antigen-binding fragment that together form the antigen-binding site of the antibody. Each variable region domain contains three CDRs, termed CDR1, CDR2, and CDR3. Accordingly, the variable heavy chain domain (VH) comprises CDR-H1, CDR-H2 and CDR-H3, and the variable light chain domain (VL) comprises CDR-L1, CDR-L2 and CDR-L3. The three CDRs are not contiguous along the linear amino acid sequence, but are close together in the folded polypeptide. The CDRs are located within loops connecting the parallel strands of the beta sheets of the variable domains.

The term "framework (FR) amino acid residues" as used herein refers to those amino acids in the framework region of the Ig chain. The term "framework region" or "FR region" as used herein includes amino acid residues that are part of the variable region but are not part of the CDRs. Thus, the variable region framework is between about 100-120 amino acids in length, but includes only those amino acids outside the CDRs.

As used herein, the term "% identical" or "percent identical" means that when comparing two sequences over a specified region, the two sequences have the specified number of identical residues at the same position. The term "% similar" or "percentage similar" has a similar meaning, but contemplates conservative substitutions in which amino acids are not identical, in addition to the number of identical amino acids between the two sequences. Known computer algorithms such as BLASTP, BLASTN and FASTA programs (Altschul, SF et al., J Mol Biol 215:403 (1990)) can be used, using, for example, Pearson et al., Proc Natl Acad Sci USA 85:2444 (1988) Default parameters are used to determine percent identity. For example, identity can be determined using the BLAST function of the National Center for Biotechnology Information (NCBI) database.

In certain embodiments, the antibodies provided herein are humanized antibodies. A "humanized antibody" is an antibody molecule that binds the desired antigen, has one or more CDRs from a non-human species (eg, a mouse antibody), and has framework regions and/or constant structures from a human immunoglobulin molecule At least some parts of the domain. Known human Ig sequences are published, for example, at ncbi.nlm.nih.gov/entrez-/query.fcgi; atcc.org/phage/hdb.html; sciquest.com; abcam.com; antibodyresource.com/onlinecomp.html; and Kabat et al., Sequences of Proteins of Immunological Interest, U.S. Dept. Health (1983). The introduced human sequences can be used to reduce immunogenicity or to reduce, enhance or modify binding affinity, on-rate, off-rate, avidity, specificity, half-life or any other suitable characteristic, as is known in the art. Art-recognized methods for antibody humanization are described in Jones et al., Nature 321:522 (1986); Verhoeyen et al., Science 239:1534 (1988); Sims et al., J Immunol 151:2296 (1993 ); Chothia and Lesk, J Mol Biol 196:901 (1987); Carter et al, Proc Natl Acad Sci USA 89:4285 (1992); Presta et al, J Immunol 151:2623 (1993); U.S. Pat. Nos. 5,589,205, 5,565,332, 6,180,370, 6,632,927, 7,241,877, 7,244,615, 7,244,832, 7,262,050; and US Patent Publication No. 2004/0236078 (filed April 30, 2004), the entire contents of which are incorporated herein by reference.

In certain embodiments, certain framework residues in the humanized antibodies provided herein have been substituted with corresponding residues from a CDR donor antibody, for example, with framework residues from a mouse anti-human CXCR3 antibody, to Altering (eg, improving) antigen binding. These framework substitutions have been identified by modeling the interactions of CDRs and framework residues to identify framework residues that are more important for antigen binding, and sequence comparisons have been performed to identify unusual framework residues at specific positions. In some embodiments, the humanized antibody variants of the disclosure are prepared using 4D humanization. See WO2009/032661 (which is incorporated herein by reference in its entirety), eg, methods for 4D humanization in paragraphs [0037]-[0044]. Briefly, 4D humanization can involve: (a) building a 3-D model of the variable domain to be humanized; (b) using molecular dynamics simulations of the 3-D model of the domain to identifying flexible residues in the variable domain; (c) identifying the closest human germline by comparing the molecular dynamics trajectories of the 3-D model to those of 49 human germlines; and (d) mutating flexible residues that are not part of the CDRs to their human germline counterparts (as identified in step (c)).

In some embodiments, humanized CXCR3 antibodies or antigen-binding fragments thereof comprising the VH and VL sequences listed in Table 1 are provided.

In some embodiments, humanized CXCR3 antibodies or antigen-binding fragments thereof comprising the heavy chain (HC) and light chain (LC) sequences listed in Table 2 are provided.

Antibody effector function/depletion activity

In certain embodiments, the anti-CXCR3 antibodies disclosed herein have the ability to direct the depletion of CXCR3-expressing cells, or can be engineered to have an enhanced ability to direct the depletion of CXCR3-expressing cells for the treatment of CXCR3-related diseases and disorders. CXCR3 expressing cells that can be depleted by the antibodies disclosed herein can include CD4 ⁺ T cells and/or CD8 ⁺ T cells. CXCR3 expressing cells that can be depleted by the antibodies disclosed herein can include CD4 ⁺ memory T cells and/or CD8 ⁺ memory T cells. As used herein, "depletion" with respect to CXCR3 ⁺ cells (ie, cells expressing CXCR3 on the cell surface) refers to the removal of these cells from a population of cells. References to depletion include full or partial depletion. Furthermore, depletion may be permanent or temporary, and may vary in magnitude and/or location. Depletion may be the result of cell death, such as apoptosis or necrosis. Antibodies can be detected by any method known in the art (e.g., flow cytometry) before and after exposure to the antibodies or antigen-binding fragments provided herein, or in the absence and presence of the antibodies or antigen-binding fragments provided herein. , immunohistochemistry, etc.) to assess depletion by measuring the number of CXCR3 ⁺ cells in the cell population. CXCR3 ⁺ cells may be depleted by at least or about 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55% following exposure to an antibody or antigen-binding fragment provided herein. %, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95% or more.

In certain embodiments, the humanized anti-human CXCR3 antibody is expressed on cells expressing human CXCR3 on the surface compared to a corresponding humanized anti-human CXCR3 antibody having a wild-type Fc region (e.g., wild-type human IgG1 Fc). Enhanced effector function. As used herein, "enhanced effector function" refers to antibody-dependent cytotoxicity ( A measurable increase in any one or more of ADCC), complement-mediated cytotoxicity (CDC), or antibody-dependent cell-mediated phagocytosis (ADCP). In certain embodiments, the reference antibody comprises a variant human Fc region. In certain embodiments, the effector function is ADCC, ADCP, or CDC, or any combination thereof. In certain embodiments, the effector function is ADCC, or CDC, or both ADCC and CDC. In certain embodiments, the effector function is ADCC. In certain embodiments, the effector function is CDC. In certain embodiments, the effector functions are ADCC and CDC. In certain embodiments, the effector function is ADCP.

As used herein, "variant human IgG1 Fc region" refers to a human IgG1 Fc region that has been engineered or modified to include one or more amino acid mutations or amino acid modifications compared to wild-type human IgG1 Fc.在某些实施方案，所述野生型人IgG1 Fc区包含氨基酸序列SLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG(EU编号192-446)(SEQ IDNO:2)。

In certain embodiments, the variant human IgGl Fc region comprises at least one of the following amino acid substitutions: G236A, S239D, S267E, H268F, S324T, I332E (Eu numbering), or any combination thereof. In certain embodiments, the variant human IgG1 Fc region comprises at least one of the following amino acid substitution groups: S239D/I332E, G236A/S267E/H268F/S324T/I332E, and S239D/H268F/S324T/I332E (Eu numbering) . In certain embodiments, the variant human IgGl Fc region comprises the amino acid substitution S239D/I332E. In other embodiments, the variant human IgGl Fc region comprises the amino acid substitutions G236A/S267E/H268F/S324T/I332E. In still other embodiments, the variant human IgGl Fc region comprises amino acid substitutions S239D/H268F/S324T/I332E.

For example, in certain embodiments, the variant human IgG1 Fc region comprises the amino acid sequence of SEQ ID NO: 3, 4, 5, 6, 7 or 8.

In certain embodiments, the variant human IgG1 Fc region comprises a sequence at least 90% identical to any one or more of SEQ ID NOs: 3-8, provided that in each case the specified amino acids are maintained replace. In various embodiments, the variant human IgG1 Fc region comprises at least 90% identical, at least 91% identical, at least 92% identical, at least 93% identical, Sequences that are at least 94% identical, at least 95% identical, at least 96% identical, at least 97% identical, at least 98% identical or at least 99% identical, provided that in each case the specified amino acid substitutions are maintained.

In certain embodiments, the variant human IgG1 Fc region comprises at least one of the following amino acid substitution groups: S239D/I332E, G236A/S267E/H268F/S324T/I332E, and S239D/H268F/S324T/I332E (Eu numbering) . For example, in certain embodiments, the variant human IgG1 Fc region comprises the amino acid sequence of SEQ ID NO:9, 10 or 11.

In certain embodiments, at least one amino acid substitution is S239D/I332E (Eu numbering). In other embodiments, at least one amino acid substitution is G236A/S267E/H268F/S324T/I332E (Eu numbering). In certain embodiments, at least one amino acid substitution is S239D/H268F/S324T/I332E (Eu numbering).

In certain embodiments, the variant human IgG1 Fc region of a humanized anti-CXCR3 antibody provided herein comprises a sequence at least 90% identical to any one of SEQ ID NOs: 9-11, provided that, in each case under maintenance of specific amino acid substitutions and enhanced effector function. In various embodiments, the variant human IgG1 Fc region comprises at least 90% identical, at least 91% identical, at least 92% identical, at least 93% identical, Sequences that are at least 94% identical, at least 95% identical, at least 96% identical, at least 97% identical, at least 98% identical or at least 99% identical, provided that in each case the specified amino acid substitutions are maintained.

CDR variants:

In addition to the foregoing embodiments, provided herein are anti-CXCR3 antibodies comprising 6 CDRs, wherein the VH comprises a CDR having the following amino acid sequence:

(i) SEQ ID NO: 12, SEQ ID NO: 35 and SEQ ID NO: 14;

(ii) SEQ ID NO: 12, SEQ ID NO: 35 and SEQ ID NO: 45;

(iii) SEQ ID NO: 12, SEQ ID NO: 36 and SEQ ID NO: 45;

(iv) SEQ ID NO: 12, SEQ ID NO: 37 and SEQ ID NO: 14;

(v) SEQ ID NO: 12, SEQ ID NO: 37 and SEQ ID NO: 45;

(vi) SEQ ID NO: 12, SEQ ID NO: 37 and SEQ ID NO: 46

(vii) SEQ ID NO: 12, SEQ ID NO: 38 and SEQ ID NO: 14;

(viii) SEQ ID NO: 12, SEQ ID NO: 38 and SEQ ID NO: 45;

(ix) SEQ ID NO: 12, SEQ ID NO: 39 and SEQ ID NO: 14;

(x) SEQ ID NO: 12, SEQ ID NO: 39 and SEQ ID NO: 47;

(xi) SEQ ID NO: 12, SEQ ID NO: 40 and SEQ ID NO: 14;

(xii) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 45

(xiii) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 46;

(xiv) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 48;

(xv) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 49;

(xvi) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 50;

(xvii) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 51;

(xviii) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 52;

(xix) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 53;

(xx) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 54;

(xxi) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 55;

(xxii) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 56;

(xxiii) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 57;

(xxiv) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 58;

(xxv) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 59;

(xxvi) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 60;

(xxvii) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 61;

(xxviii) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 62;

(xxix) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 63;

(xxx) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 64;

(xxxi) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 65;

(xxxii) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 66;

(xxviii) SEQ ID NO:34, SEQ ID NO:13 and SEQ ID NO:14;

(xxxiv) SEQ ID NO: 12, SEQ ID NO: 41 and SEQ ID NO: 14;

(xxxv) SEQ ID NO: 12, SEQ ID NO: 41 and SEQ ID NO: 46;

(xxxvi) SEQ ID NO: 12, SEQ ID NO: 42 and SEQ ID NO: 14;

(xxxvi) SEQ ID NO: 12, SEQ ID NO: 42 and SEQ ID NO: 45;

(xxxviii) SEQ ID NO: 12, SEQ ID NO: 43 and SEQ ID NO: 14;

(xxxix) SEQ ID NO: 12, SEQ ID NO: 44 and SEQ ID NO: 14;

(xl) SEQ ID NO: 12, SEQ ID NO: 41 and SEQ ID NO: 14; or

(xli) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 68; and

Wherein said VL comprises a CDR having the following amino acid sequence:

(i) SEQ ID NO:69, SEQ ID NO:16 and SEQ ID NO:17;

(ii) SEQ ID NO:70, SEQ ID NO:16 and SEQ ID NO:17;

(iii) SEQ ID NO:71, SEQ ID NO:16 and SEQ ID NO:17;

(iv) SEQ ID NO:71, SEQ ID NO:16 and SEQ ID NO:75;

(v) SEQ ID NO:72, SEQ ID NO:16 and SEQ ID NO:17;

(vi) SEQ ID NO:73, SEQ ID NO:16 and SEQ ID NO:17;

(vii) SEQ ID NO: 74, SEQ ID NO: 16 and SEQ ID NO: 17;

(viii) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 75;

(ix) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 76;

(x) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 77;

(xi) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 78; or

(vii) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 79.

In addition to the foregoing embodiments, provided herein are anti-CXCR3 antibodies comprising 6 CDRs, wherein the VH comprises a CDR having the following amino acid sequence:

(i) SEQ ID NO: 12, SEQ ID NO: 35 and SEQ ID NO: 14;

(ii) SEQ ID NO: 12, SEQ ID NO: 35 and SEQ ID NO: 45;

(iii) SEQ ID NO: 12, SEQ ID NO: 36 and SEQ ID NO: 45;

(iv) SEQ ID NO: 12, SEQ ID NO: 37 and SEQ ID NO: 14;

(v) SEQ ID NO: 12, SEQ ID NO: 37 and SEQ ID NO: 45;

(vi) SEQ ID NO: 12, SEQ ID NO: 37 and SEQ ID NO: 46;

(vii) SEQ ID NO: 12, SEQ ID NO: 38 and SEQ ID NO: 14;

(viii) SEQ ID NO: 12, SEQ ID NO: 38 and SEQ ID NO: 45;

(ix) SEQ ID NO: 12, SEQ ID NO: 39 and SEQ ID NO: 14;

(x) SEQ ID NO: 12, SEQ ID NO: 39 and SEQ ID NO: 47; or

(xi) SEQ ID NO: 12, SEQ ID NO: 40, and SEQ ID NO: 14; and

Wherein said VL comprises a CDR having the following amino acid sequence:

(i) SEQ ID NO:69, SEQ ID NO:16 and SEQ ID NO:17;

(ii) SEQ ID NO:70, SEQ ID NO:16 and SEQ ID NO:17;

(iii) SEQ ID NO:71, SEQ ID NO:16 and SEQ ID NO:17;

(iv) SEQ ID NO:71, SEQ ID NO:16 and SEQ ID NO:100;

(v) SEQ ID NO:72, SEQ ID NO:16 and SEQ ID NO:17;

(vi) SEQ ID NO:73, SEQ ID NO:16 and SEQ ID NO:17;

(vii) SEQ ID NO: 74, SEQ ID NO: 16 and SEQ ID NO: 17;

(viii) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 75;

(ix) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 76;

(x) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 77;

(xi) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 78; or

(vii) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 79.

In addition to the foregoing embodiments, provided herein are anti-CXCR3 antibodies comprising 6 CDRs, wherein the VH comprises a CDR having the following amino acid sequence:

(i) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 45;

(ii) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 46;

(iii) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 48;

(iv) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 49;

(v) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 50;

(vi) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 51;

(vii) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 52;

(viii) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 53;

(ix) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 54;

(x) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 55; or

(xi) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 56; and

Wherein said VL comprises a CDR having the following amino acid sequence:

(i) SEQ ID NO:69, SEQ ID NO:16 and SEQ ID NO:17;

(ii) SEQ ID NO:70, SEQ ID NO:16 and SEQ ID NO:17;

(iii) SEQ ID NO:71, SEQ ID NO:16 and SEQ ID NO:17;

(iv) SEQ ID NO:71, SEQ ID NO:16 and SEQ ID NO:75;

(v) SEQ ID NO:72, SEQ ID NO:16 and SEQ ID NO:17;

(vi) SEQ ID NO:73, SEQ ID NO:16 and SEQ ID NO:17;

(vii) SEQ ID NO: 74, SEQ ID NO: 16 and SEQ ID NO: 17;

(viii) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 75;

(ix) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 76;

(x) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 77;

(xi) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 78; or

(vii) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 79.

In addition to the foregoing embodiments, provided herein are anti-CXCR3 antibodies comprising 6 CDRs, wherein the VH comprises a CDR having the following amino acid sequence:

(i) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 57;

(ii) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 58;

(iii) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 59;

(iv) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 60;

(v) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 61;

(vi) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 62;

(vii) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 63;

(viii) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 64;

(ix) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 65; or

(x) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 66; and

Wherein said VL comprises a CDR having the following amino acid sequence:

(i) SEQ ID NO:69, SEQ ID NO:16 and SEQ ID NO:17;

(ii) SEQ ID NO:70, SEQ ID NO:16 and SEQ ID NO:17;

(iii) SEQ ID NO:71, SEQ ID NO:16 and SEQ ID NO:17;

(iv) SEQ ID NO:71, SEQ ID NO:16 and SEQ ID NO:75;

(v) SEQ ID NO:72, SEQ ID NO:16 and SEQ ID NO:17;

(vi) SEQ ID NO:73, SEQ ID NO:16 and SEQ ID NO:17;

(vii) SEQ ID NO: 74, SEQ ID NO: 16 and SEQ ID NO: 17;

(viii) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 75;

(ix) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 76;

(x) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 77;

(xi) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 78; or

(vii) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 79.

In addition to the foregoing embodiments, provided herein are anti-CXCR3 antibodies comprising 6 CDRs, wherein the VH comprises a CDR having the following amino acid sequence:

(i) SEQ ID NO:34, SEQ ID NO:13 and SEQ ID NO:14;

(ii) SEQ ID NO: 12, SEQ ID NO: 41 and SEQ ID NO: 14;

(iii) SEQ ID NO: 12, SEQ ID NO: 41 and SEQ ID NO: 71;

(iv) SEQ ID NO: 12, SEQ ID NO: 42 and SEQ ID NO: 14;

(v) SEQ ID NO: 12, SEQ ID NO: 42 and SEQ ID NO: 70;

(vi) SEQ ID NO: 12, SEQ ID NO: 43 and SEQ ID NO: 14;

(vii) SEQ ID NO: 12, SEQ ID NO: 44 and SEQ ID NO: 14;

(viii) SEQ ID NO: 12, SEQ ID NO: 41 and SEQ ID NO: 14; or

(ix) SEQ ID NO: 12, SEQ ID NO: 13 and SEQ ID NO: 68; and

Wherein said VL comprises a CDR having the following amino acid sequence:

(i) SEQ ID NO:69, SEQ ID NO:16 and SEQ ID NO:17;

(ii) SEQ ID NO:70, SEQ ID NO:16 and SEQ ID NO:17;

(iii) SEQ ID NO:71, SEQ ID NO:16 and SEQ ID NO:17;

(iv) SEQ ID NO:71, SEQ ID NO:16 and SEQ ID NO:75;

(v) SEQ ID NO:72, SEQ ID NO:16 and SEQ ID NO:17;

(vi) SEQ ID NO:73, SEQ ID NO:16 and SEQ ID NO:17;

(vii) SEQ ID NO: 74, SEQ ID NO: 16 and SEQ ID NO: 17;

(viii) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 75;

(ix) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 76;

(x) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 77;

(xi) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 78; or

(vii) SEQ ID NO: 15, SEQ ID NO: 16 and SEQ ID NO: 79.

In addition to the foregoing embodiments, provided herein are anti-CXCR3 antibodies comprising a VH comprising the following amino acid sequence:

SEQ ID NO:80, SEQ ID NO:81, SEQ ID NO:82, 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 or SEQ ID NO:90.

In addition to the foregoing embodiments, provided herein are anti-CXCR3 antibodies comprising a VH comprising the following amino acid sequence:

SEQ ID NO:91, SEQ ID NO:92, SEQ ID NO: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, SEQ ID NO:100 or SEQ ID NO:101.

In addition to the foregoing embodiments, provided herein are anti-CXCR3 antibodies comprising a VH comprising an amino acid sequence selected from the group consisting of:

SEQ ID NO: 102, SEQ ID NO: 103, SEQ ID NO: 104, SEQ ID NO: 105,

SEQ ID NO: 106, SEQ ID NO: 107, SEQ ID NO: 108, SEQ ID NO: 109,

SEQ ID NO: 110 or SEQ ID NO: 111.

In addition to the foregoing embodiments, provided herein are anti-CXCR3 antibodies comprising a VH comprising the following amino acid sequence:

SEQ ID NO: 112, SEQ ID NO: 113, SEQ ID NO: 114, SEQ ID NO: 115,

SEQ ID NO: 116, SEQ ID NO: 117, SEQ ID NO: 118, SEQ ID NO: 119,

SEQ ID NO: 120, SEQ ID NO: 121 or SEQ ID NO: 122.

In addition to the foregoing embodiments, provided herein are anti-CXCR3 antibodies comprising a VL comprising the following amino acid sequence:

SEQ ID NO: 123, SEQ ID NO: 124, SEQ ID NO: 125, SEQ ID NO: 126,

SEQ ID NO: 127, SEQ ID NO: 128, SEQ ID NO: 129, SEQ ID NO: 130,

SEQ ID NO:131, SEQ ID NO:132, SEQ ID NO:133 or SEQ ID NO:134.

In certain embodiments, an anti-human CXCR3 antibody comprises a VH/VL pair comprising the amino acid sequences shown in Table 3, respectively (indicated changes are relative to the corresponding 53Hu37 VH or VL sequences, respectively (SEQ ID NO: 20 and 24) ):

table 3

neutralizing antibody

In certain embodiments, the humanized anti-human CXCR3 antibodies provided herein are CXCR3 neutralizing antibodies. In certain exemplary embodiments, the CXCR3 antibody has neutralizing activity in addition to enhanced effector function. The combined effects of CXCR3 neutralization and CXCR3 ⁺ cell depletion may be advantageous whenever it is desired to reduce or eliminate a CXCR3-mediated effect, such as recruitment of T cells.

A "CXCR3 neutralizing antibody" binds to CXCR3 and blocks the activity of the receptor, such as the typical physiological and genetic responses resulting from the binding of a CXCR3 ligand to CXCR3. The neutralizing activity can be complete (100% neutralization) or partial (e.g., about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95% (or between (any percentage) or more) and will depend on various factors known to the skilled artisan, such as antibody concentration, affinity, and epitope, and the particular assay used to assess neutralizing activity. The neutralizing activity of a CXCR3 neutralizing antibody can be shown by assays to measure inhibition of, eg, ligand binding, GTP binding, calcium mobilization, cell chemotaxis and/or receptor internalization. Many assays for determining the activity of neutralizing antibodies, in particular CXCR3 neutralizing antibodies, are known to the skilled person and can be readily adapted to verify whether a particular antibody is neutralizing.

For example, in some embodiments, the neutralizing activity of an anti-CXCR3 antibody can be assessed by a chemotaxis assay substantially as described in the package insert for the antibody produced by clone 49801 and sold by R&D Systems (Cat. No.: MAB160) . The neutralizing dose- ₅₀ (ND50) was defined as the concentration of antibody required to produce a half-maximal inhibition of cell surface CXCR3-mediated recombinant human CXCL11 (rhCXCL11) responses in responding cell lines at a specific rhCXCL11 concentration. To measure the ability of antibodies to block rhCXCL11-induced chemotaxis of hCXCR3-transfected BaF/3 cells, 7 ng/mL of rhCXCL11 was added to the lower compartment of a 96-well chemotaxis chamber (NeuroProbe, Cabin John, Md.). Chemotaxis chambers were then assembled using PVP-free polycarbonate filters (5 μm pore size). Add serial dilutions of antibody (eg, 0.001 to 10000 μg/mL) and ^0.25x106 cells/well to the top well of the chemotaxis chamber. After incubation for 3 hours at 37 °C in a 5% _CO2 humidified incubator, disassemble the chemotaxis chamber, transfer the cells that migrated to the lower chamber to a working plate, and quantify using, for example, Resasurin Fluorescence.

Colvin et al., Mol Cell Biol 26:5838-49 (2006) describe other assays that can be used in certain embodiments to determine the neutralizing activity of neutralizing anti-CXCR3 antibodies. Briefly, 300-19 cells, a murine pre-B-cell leukemia cell line that functionally expresses CXCR4, can be used. Following transfection, this cell line can functionally express other chemokine receptors, such as human CXCR3 (see, eg, paragraphs 201-209 of US Patent Application Publication No. 2010/0061983, which is incorporated herein by reference). 300-19 cells expressing human CXCR3 can be grown in complete RPMI medium containing 10% fetal bovine serum (FBS). To assess the binding of CXCR3 ligands to CXCR3 in the presence of candidate neutralizing R3 antibodies, 400,000 CXCR3/300-19 cells were plated in 96-well tissue culture plates in a total volume of 150 μL of binding buffer (0.5% BSA, 5 mM MgCl ₂ , 1 mM CaCl ₂ , 50 mM HEPES, pH 7.4). A total of 0.04 nM of ¹²⁵ I-labeled CXCL10 (New England Nuclear, Boston, Mass.) or CXCL11 (Amersham Biosciences Piscataway, NJ) and 5×10 ⁶ nM to 500 nM of unlabeled CXCL10 or CXCL11 (Peprotech, Rocky Hill, NJ) can be added. Add to cells and incubate with shaking for 90 minutes at room temperature. Cells were transferred to 96-well filter plates (Millipore, Billerica, Mass.) presoaked in 0.3% polyethyleneimine and washed three times with 200 μL of binding buffer supplemented with 0.5 M NaCl. Plates were dried and radioactivity was measured after addition of scintillation fluid in a Wallac Microbeta scintillation counter (Perkin-Elmer Life Sciences, Boston, Mass.). Binding of CXCL9 can be assessed similarly to CXCL10 and CXCL11.

In certain embodiments, the antibodies disclosed herein prevent or reduce calcium flux into CXCR3 expressing cells. In some embodiments, calcium flux can be detected in cells such as CXCR3/300-19 cells. Approximately 5 x 10 ⁶ cells were suspended in 2 mL of RPMI medium containing 1% bovine serum albumin (BSA). 15 micrograms of Fura-2 (Molecular Probes, Eugene, OR) was added and the cells were incubated at 37°C for 20 minutes. Cells were washed twice in PBS and resuspended in 2 mL of calcium flux buffer (145 mM NaCl, 4 mM KCl, 1 mM NaHPO ₄ , 1.8 mM CaCl ₂ , 25 mM HEPES, 0.8 mM MgCl ₂ and 22 mM glucose). Fluorescence readings were measured on a DeltaRAM Fluorometer (Photon Technology International, Lawrenceville, NJ) at 37°C. Before and after the addition of chemokines (e.g., CXCL9, CXCL10, or CXCL11), intracellular calcium concentrations were recorded as excitation fluorescence intensity emitted at 510 nm in response to sequential excitation at 340 nm and 380 nm, and expressed as 340 nm versus 380 nm. The relative ratio of fluorescence at .

In certain embodiments, CXCR3 neutralization can be assessed by measuring a reduction in receptor internalization. In some embodiments, about 2.5 x 10 ⁵ cells (such as CXCR3/300-19 cells) can be incubated at 37°C with different concentrations of CXCL10, CXCL11 or CXCL9 in RPMI medium containing 1% BSA. Incubate for 30 minutes for receptor internalization assay. Cells can then be washed with ice-cold flow cytometry staining buffer, followed by analysis of surface expression of CXCR3 using a PE-conjugated CXCR3 antibody.

In certain embodiments, neutralizing anti-CXCR3 antibodies may have a concentration of about 0.01, 0.02, 0.05, 0.1, 0.2, 0.5, 1, 2, 3, 4, 5, 6, 7, as assessed by any of the above assays. ND50 of 8, 9, 10, 15, 20, 40, ₅₀ or 100 μg/mL. In particular embodiments, the _ND50 may be 0.5-12 μg/mL, and in more particular embodiments, 1-6 μg/mL.

Inhibition of cell migration, recruitment, or accumulation by the antibodies or antigen-binding fragments provided herein can be assessed by any method known to those of skill in the art. These methods may include, for example, analysis of biopsies by immunohistochemistry, flow cytometry, RT-PCR, etc., to assess one or more cell populations or cells at one or more locations within the body or organ ( such as the number of CXCR3 ⁺ cells). Cell migration, recruitment or accumulation can be inhibited by at least or about 10%, 15%, 20%, 25%, 30% compared to cell migration, recruitment or accumulation in the absence of an antibody or antigen-binding fragment provided herein , 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95% or more.

Nucleotide sequence

Also provided herein are nucleotide sequences encoding the amino acid sequences disclosed herein. In some embodiments, the nucleotide sequence encodes an antibody or fragment capable of depleting CXCR3 ⁺ cells in vitro and/or in vivo. In certain embodiments, the nucleotide sequences can be used to prepare expression vectors for expression of anti-CXCR3 antibodies or antigen-binding fragments thereof in cells (eg, expression in mammalian cells).

In certain embodiments, also disclosed herein are polynucleotides substantially identical to those encoding the amino acid sequences disclosed herein. Substantially identical sequences may be polymorphic sequences, ie alternative sequences or alleles in a population. Substantially identical sequences may also contain mutagenic sequences, including sequences containing silent mutations. A mutation may comprise a change of one or more nucleotide residues, a deletion of one or more nucleotide residues, or an insertion of one or more additional nucleotide residues. Due to the degeneracy of nucleic acid encoding, substantially identical sequences may also encompass various nucleotide sequences encoding the same amino acid at any given amino acid position in the amino acid sequences disclosed herein. Also included within the substantially identical sequences are sequences encoding one or more chains of antibodies that retain the ability to deplete CXCR3 ⁺ cells in vitro and/or in vivo.

In certain embodiments, a nucleic acid provided herein encodes the amino acid sequence of one or more chains of an antibody or fragment provided herein that is capable of depleting CXCR3 expressing cells, or the nucleic acid can be expressed under stringent conditions Hybridizes to nucleic acid encoding the amino acid sequence of one or more chains of the antibody or antigen-binding fragment thereof.

In certain embodiments, disclosed herein is a polynucleotide sequence comprising a nucleotide sequence encoding the amino acid sequence of the VH domain of an anti-CXCR3 antibody or antigen-binding fragment thereof, combined with a core encoding the heavy chain of said antibody The nucleotide sequence is at least about 80%-100% (e.g., about 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% (or any percentage in between)) the same. In certain embodiments, the polynucleotide sequence may comprise 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, or Nucleotide sequences of 10 mutations (including additions, deletions and substitutions, such as conservative substitutions).

In certain embodiments, disclosed herein are polynucleotide sequences comprising a nucleotide sequence encoding the amino acid sequence of the VL domain of an anti-CXCR3 antibody or fragment, in combination with a nucleotide sequence encoding the light chain of the antibody At least about 80%-100% (e.g., about 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% ( or any percentage in between)) the same. In certain embodiments, the polynucleotide sequence may comprise 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, or Nucleotide sequences of 10 mutations (including additions, deletions and substitutions, such as conservative substitutions).

In particular embodiments, the polynucleotide sequences disclosed herein comprise at least about 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97% of the VH amino acid sequence , 98%, 99%, or 100% (or any percentage in between) identical nucleotide sequence, or at least about 80%, 85%, 90%, 91%, 92%, 93%, 94% identical to the VL amino acid sequence %, 95%, 96%, 97%, 98%, 99%, or 100% (or any percentage in between) identical nucleotide sequences, wherein the nucleotide sequences encode the heavy chains of any of the antibodies disclosed herein and light chain amino acid sequences.

The disclosed polynucleotides can be obtained by any method known in the art. For example, if the nucleotide sequence of an antibody is known, a polynucleotide encoding the antibody can be assembled from chemically synthesized oligonucleotides. This may involve, for example, synthesizing overlapping oligonucleotides containing portions of the sequence encoding the antibody, annealing and ligating those oligonucleotides, and then amplifying the ligated oligonucleotides by PCR. The disclosed polynucleotides can also be generated from any other suitable nucleic acid source, such as an antibody cDNA library, or cDNA isolated from any tissue or cell that expresses the antibody (e.g., from a hybridoma cell selected to express the antibody) library.

Expression of anti-CXCR3 antibodies or antigen-binding fragments thereof

Following manipulation of nucleic acid encoding a humanized anti-CXCR3 antibody or antigen-binding fragment thereof provided herein, the encoding nucleic acid is typically inserted into an expression vector for introduction into a host cell that can be used to produce the desired amount of the encoded antibody or its antigen-binding fragment. Antigen-binding fragments. Suitable vectors for expression are known in the art. Suitable host cells include, for example, CHO, COS, Sf9 and/or other human or non-human cell lines. In some embodiments, host cells transiently or stably express nucleic acids on vectors when cultured in culture, thereby providing methods for producing the antibodies or fragments disclosed herein.

The term "vector" or "expression vector" as used herein describes a vehicle for introducing and expressing a desired gene in a cell. Such vectors include, for example, plasmids, bacteriophage, viruses and retroviruses, as known to those skilled in the art. In general, suitable vectors will contain a selectable marker, appropriate restriction sites to facilitate cloning of the desired gene, and the ability to enter and/or replicate in eukaryotic or prokaryotic cells.

A wide variety of expression vector systems can be employed for expressing the anti-CXCR3 antibodies provided herein. For example, one class of vectors utilizes DNA elements derived from animal viruses such as bovine papilloma virus, polyoma virus, adenovirus, vaccinia virus, baculovirus, retrovirus (RSV, MMTV or MOMLV) or SV40 virus. Other vector classes involve the use of polycistronic systems with internal ribosome binding sites. Additionally, cells that have integrated DNA into their chromosomes can be selected by introducing one or more markers that allow selection of transfected host cells. The markers may confer prototrophy, biocide resistance (eg antibiotics) or resistance to heavy metals such as copper in an auxotrophic host. A selectable marker gene can be directly linked to the DNA sequence to be expressed, or introduced into the same cell by co-transformation. Additional elements may also be required for optimal mRNA synthesis. These elements may include signal sequences, splicing signals, and transcriptional promoters, enhancers, and termination signals. In some embodiments, the cloned variable region genes are inserted into an expression vector along with the heavy and light chain constant region genes as described above. In some embodiments, the heavy and light chain constant regions are human.

In other embodiments, polycistronic constructs can be used to express the anti-CXCR3 antibodies or antigen-binding fragments thereof provided herein. In such expression systems, multiple gene products of interest, such as the heavy and light chains of antibodies, can be produced from a single polycistronic construct. These systems advantageously use internal ribosome entry sites (IRES) to provide relatively high levels of the polypeptides provided herein in eukaryotic host cells. Compatible IRES sequences are disclosed in US Patent No. 6,193,980, which is incorporated herein by reference. Those skilled in the art will appreciate that such expression systems can be used to efficiently produce the repertoire of polypeptides disclosed in this application.

Once a vector or DNA sequence encoding an antibody or antigen-binding fragment thereof has been prepared, the expression vector can be introduced into a suitable host cell. That is, the host cell can be transformed. Introduction of plasmids into host cells can be accomplished by various techniques well known to those skilled in the art. These techniques include, but are not limited to, transfection (including electrophoresis and electroporation), protoplast fusion, calcium phosphate precipitation, cell fusion with envelope DNA, microinjection, and whole virus infection. See Ridgway, A.A.G. "Mammalian Expression Vectors" Chapter 24.2, pp. 470-472 Vectors, Rodriguez and Denhardt, eds. (Butterworths, Boston, Mass. 1988). In some embodiments, the plasmid is introduced into the host by electroporation. Transformed cells are grown under conditions suitable for the production of the encoding amino acid sequences (eg, antibody light and heavy chains), and these cells are assayed for production of the encoding amino acid sequences. Exemplary assay techniques include enzyme-linked immunosorbent assay (ELISA), radioimmunoassay (RIA), flow cytometry, immunohistochemistry, and the like.

A "host cell" refers to a cell into which has been introduced a vector constructed using recombinant nucleic acid techniques and encoding at least one foreign protein. In describing processes for isolating polypeptides from recombinant hosts, the terms "cell" and "cell culture" are used interchangeably to denote the source of an encoded protein (eg, an antibody or antigen-binding fragment thereof), unless expressly stated otherwise. In other words, recovery of a polypeptide from "cells" may mean recovery from whole cells pelleted by centrifugation, or from cell cultures containing both media and suspension cells.

In one embodiment, the host cell line used for antibody expression is of mammalian origin; one skilled in the art can determine the particular host cell line most suitable for expression of the desired gene product therein. Exemplary host cell lines include, but are not limited to, DG44 and DUXB11 (Chinese hamster ovary line, DHFR-), HELA (human cervical carcinoma), CVI (monkey kidney line), COS (derivative of CVI with SV40 T antigen), R1610 (Chinese hamster fibroblasts), BALBC/3T3 (mouse fibroblasts), SP2/O (mouse myeloma), BFA-1c1BPT (bovine endothelial cells), RAJI (human lymphocytes), 293 (human kidney) . In one embodiment, NSO cells can be used. In some embodiments, CHO cells are used. Host cell lines are typically available from commercial services, the American Tissue Culture Collection or from the published literature.

In one embodiment, the cell line provides altered glycosylation, e.g., afucosylation, of the antibody expressed therefrom (e.g., PER.C6.RTM (Crucell) or FUT8 knockout CHO cell line (Potelligent. RTM. Cells) (Biowa, Princeton, NJ)). Alternatively, cells may lack one or more glycosidases required for early processing of N-glycans and/or culture conditions may inhibit the activity of one or more of these glycosidases. For example, a cell may be deficient in one or more glycosidases, such as α-glucosidase I, α-glucosidase II, and α-mannosidase I. Additionally, or alternatively, the engineered cell can be contacted with an inhibitor of one or more glycosidases (eg, α-glucosidase I, α-glucosidase II, and α-mannosidase I). In certain embodiments, the inhibitor is an inhibitor of alpha-mannosidase I, eg, a specific inhibitor of alpha-mannosidase I, a Kiff base. Exemplary methods of culturing host cells with Kiff bases and other inhibitors are disclosed in US Patent No. 8,071,336, which is incorporated herein by reference in its entirety. In certain embodiments, Kieff base treatment produces antibodies with at least 50% Man _5-9 (GlcNAc) ₂ N-glycans, with Man8 and Man9 containing N-glycans together being the predominant species.

In vitro production allows for scale-up to give large quantities of the desired polypeptide. Techniques for mammalian cell culture under tissue culture conditions are known in the art and include homogenous suspension culture (e.g. in airlift or continuous stirred reactors), or bead or ceramic on agarose Immobilized or embedded cell culture on cassettes (eg in hollow fibers, microcapsules). Solutions of polypeptides may be purified, if necessary and/or desired, by conventional chromatographic methods, eg gel filtration, ion exchange chromatography, chromatography on DEAE-cellulose and/or (immuno)affinity chromatography.

Nucleic acids encoding the anti-CXCR3 antibodies or fragments thereof provided herein can also be expressed in non-mammalian cells such as bacterial or yeast or plant cells. In this regard, it is understood that various unicellular non-mammalian microorganisms (such as bacteria) can also be used to express the antibodies and antigen-binding fragments thereof provided herein, ie, those capable of growth in culture or fermentation. Suitable bacteria include members of the family Enterobacteriaceae, such as Escherichia coli or strains of Salmonella; Bacillus family, such as Bacillus subtilis; pneumococcus; Streptococcus and Haemophilus influenzae. It is also understood that when expressed in bacteria, polypeptides can be part of inclusion bodies. Peptides must be isolated, purified, and then assembled into functional molecules.

In addition to prokaryotes, eukaryotic microorganisms can also be used. Saccharomyces cerevisiae or common baker's yeast is the most commonly used eukaryotic microorganism, although many other strains are commonly available. For expression in Saccharomyces, for example the plasmid YRp7 is commonly used (Stinchcomb et al., Nature, 282:39 (1979); Kingsman et al., Gene, 7:141 (1979); Tschemper et al., Gene, 10:157 (1980)) . This plasmid already contains the TRP1 gene which provides a selection marker for a mutant strain of yeast lacking the ability to grow in tryptophan, eg ATCC No. 44076 or PEP4-1 (Jones, Genetics, 85:12 (1977)). The presence of Trpl lesions as a genomic feature of yeast host cells then provides an efficient environment for detection of transformation by growth in the absence of tryptophan.

Pharmaceutical formulations and methods of administration

Provided herein are pharmaceutical compositions comprising a humanized anti-human CXCR3 antibody disclosed herein and a pharmaceutically acceptable carrier.

Methods of making and administering to a subject the antibodies or antigen-binding fragments thereof provided herein are well known or readily determined by those skilled in the art. The route of administration of the antibody or fragment thereof is oral, parenteral (eg, intravenous, intramuscular, intraperitoneal or subcutaneous), inhalation or topical administration. In some embodiments, the antibodies provided herein are formulated for intravenous administration. In some embodiments, suitable pharmaceutical compositions for injection comprise a buffer (eg, acetate, phosphate, or citrate buffer), a surfactant (eg, polysorbate), optionally a stabilizer (such as human albumin), etc.

Pharmaceutical compositions suitable for injectable use include sterile aqueous solutions (water soluble) or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersion. The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (for example, glycerol, propylene glycol, and liquid polyethylene glycol, etc.), and suitable mixtures thereof. Proper fluidity can be maintained, for example, by the use of coatings such as lecithin, by maintaining the desired particle size in the case of dispersion, and by the use of surfactants. Prevention of the action of microorganisms can be achieved by various antibacterial and antifungal agents (eg, parabens, chlorobutanol, phenol, ascorbic acid, thimerosal, and the like). In many cases it will be preferable to include isotonic agents, for example sugars, polyalcohols (eg mannitol, sorbitol) or sodium chloride in the compositions. Prolonged absorption of the injectable compositions can be brought about by including in the composition an agent which delays absorption, for example, aluminum monostearate and gelatin.

In any case, sterile injectable solutions can be prepared by incorporating the active compound (for example, the antibody itself or in combination with other active agents) in the required amount in an appropriate solvent, followed by filtered sterilization, according to It is desirable to have one or a combination of ingredients listed herein. Generally, dispersions are prepared by incorporating the active compound into a sterile vehicle that contains a basic dispersion medium and the required other ingredients from those enumerated above. In the case of sterile powders for the preparation of sterile injectable solutions, the preferred methods of preparation are vacuum drying and freeze-drying which yield the active ingredient plus any additional desired ingredient from a previously sterile-filtered solution thereof. of powder. Formulations for injection are processed, filled into containers such as ampoules, bags, bottles, syringes or vials, and sealed under aseptic conditions according to methods known in the art. Such articles of manufacture preferably have a label or package insert indicating that the relevant composition is useful for treating a subject suffering from or susceptible to an autoimmune or neoplastic disorder.

Dosages of the antibodies or antigen-binding fragments thereof provided herein for use in the treatment of the disorders described above vary depending on many different factors, including the mode of administration, the site of the target, the physiological state of the patient, whether the patient is human or animal, other drugs administered, and Whether treatment is prophylactic or curative. Typically, the patient is a human, but non-human mammals, including transgenic mammals, can also be treated.

In some embodiments, the dosage may range, for example, from about 0.0001 to 100 mg/kg, or 0.01 to 5 mg/kg of host body weight.

Such doses may be administered to a subject daily, every other day, weekly, or on any other schedule determined by empirical analysis. Antibodies provided herein, or antigen-binding fragments thereof, can be administered multiple times. Intervals between single administrations can be, for example, daily, weekly, monthly or yearly. Intervals may also be irregular as indicated by measuring blood polypeptide or target molecule levels in the patient.

The antibodies or antigen-binding fragments thereof provided herein can optionally be administered in combination with other agents for the treatment of a disorder or condition in need of treatment (eg, prophylactic or therapeutic treatment). Preferred additional agents are those recognized in the art and administered against the criteria of the particular disorder.

Methods of treating CXCR3-associated diseases or disorders

The CXCR3 antibodies or antigen-binding fragments thereof provided herein can be used to antagonize CXCR3 activity. In some embodiments, antibodies and antigen-binding fragments are used in methods for inhibiting the binding of CXCR3 to one or more ligands (such as CXCL9, CXCL10, and/or CXCL11); inhibiting the migration of CXCR3 ⁺ cells , accumulation, recruitment, or infiltration (eg, to sites of inflammation); and/or depletion of CXCR3 ⁺ cells. In some embodiments, the antibodies and antigen-binding fragments are used in methods of depleting CXCR3 ⁺ cells in vivo. CXCR3 ⁺ cells include, but are not limited to, CXCR3 ⁺ /CD4 ⁺ T cells, CXCR3 ⁺ /CD8 ⁺ T cells, and CXCR3 ⁺ /CD19 ⁺ B cell subsets.

In certain embodiments, methods of treating a CXCR3-associated disease or disorder by administering to a subject in need thereof a pharmaceutical composition comprising one or more CXCR3 antibodies or antigen-binding fragments thereof are provided. In one embodiment, a method of treating or reducing the progression of a T cell-mediated autoimmune disease is provided. The method comprises the step of administering a humanized anti-human CXCR3 antibody or antigen-binding fragment thereof disclosed herein to a subject in need thereof, thereby treating or reducing the progression of a T cell-mediated autoimmune disease. In certain embodiments, the T cell-mediated autoimmune disease is vitiligo. In some embodiments, subjects in need thereof include those who have been diagnosed with a CXCR3-related disease or a T-cell-mediated autoimmune disease, or are predisposed to develop a CXCR3-related disease or a T-cell-mediated autoimmune disease as described herein. subjects with autoimmune diseases.

Subjects to be treated by the methods provided herein can include humans or other mammals. In one embodiment, the subject is a human. In various embodiments, the subject may be treated prophylactically or after the onset of any disorder associated with aberrant CXCR3 activity or any disorder in which disruption of CXCR3 signaling would be therapeutically beneficial.

In some embodiments, the subject may be treated prophylactically, or treated after the onset of vitiligo.

A "subject with vitiligo" is any subject with patchy skin depigmentation, usually of autoimmune or unknown cause, consistent with a medical diagnosis of vitiligo. Affected areas may include the face, hands, feet, arms, legs, lips, or any combination thereof, but may involve other areas such as the neck or torso. In some cases, hair pigmentation is reduced.

Example

The antibodies, compositions of matter and methods are further illustrated by the following examples, which examples should not be construed as further limiting. The contents of the Sequence Listing, Figures and all references, patents and published patent applications cited throughout this application are expressly incorporated herein by reference.

Example 1: Generation of Humanized Anti-CXCR3 Monoclonal Antibody

Anti-CXCR3 monoclonal antibodies were generated as described in WO2013/109974. Humanized clone 53 monoclonal antibody was generated as described below.

A humanized version of clone 53 capable of directing depletion of CXCR3 expressing cells was prepared. A humanized clone 53 monoclonal antibody with the VH and VK sequences shown in Table 4 was generated.

Germinal Index scores for each humanized variant shown in Table 4. The heavy chain was compared to the IGHV3-23*03/IGHJ4*03 germline sequence; the light chain was compared to the IGKV1-9*01/IGKJ2*01 germline sequence.

Table 4

Hu antibody VH SEQ ID NO VL SEQ ID NO Hair growth index 1^a Hair growth index 2^b twenty three 18 twenty one 0.885 0.950 twenty four 19 twenty one 0.872 0.933 25 20 twenty one 0.877 0.939 26 18 twenty two 0.890 0.955 27 19 twenty two 0.877 0.939 29 20 twenty two 0.881 0.944 30 18 twenty three 0.890 0.955 31 19 twenty three 0.877 0.939 33 20 twenty three 0.881 0.944 34 18 twenty four 0.895 0.961 35 19 twenty four 0.881 0.944 37 20 twenty four 0.886 0.950

^a Germinative index (1) re: pairwise comparison of all residues except those occupied by the D region

light: pairwise comparison of all residues

^b Germinative ability index (2) re: only pairwise comparisons of all framework residues (as defined by the IMTG definition)

Light: only pairwise comparisons of all framework residues (as defined by the IMTG definition)

The binding characteristics of each of the humanized monoclonal antibodies provided herein are shown in Table 5.

Table 5 Binding Kinetics

Hu antibody ka(1/Ms) kd(1/s) KD(M) twenty three 7.56E+04 2.45E-05 3.22E-10 twenty four 1.01E+05 6.16E-06 6.14E-11 25 9.95E+04 6.21E-05 6.23E-10 26 1.01E+05 5.46E-05 5.37E-10 27 1.13E+05 8.34E-06 7.37E-11 29 1.15E+05 3.50E-05 3.03E-10 30 9.37E+04 1.26E-04 1.34E-09 31 1.11E+05 5.00E-05 4.49E-10 33 9.66E+04 6.35E-05 6.56E-10 34 8.89E+04 9.19E-05 1.03E-09 35 1.02E+05 4.73E-06 4.56E-11 37 6.74E+04 6.48E-05 9.57E-10

Table 6 shows the binding characteristics and germinal capacity of clone 53 (53), chimeric clone 53 (Ch53) and humanized versions of clone 53 (53Hu1-53Hu20)

Table 6

^a Germinative index (1) re: pairwise comparison of all residues except those occupied by the D region

light: pairwise comparison of all residues

^b Germinative ability index (2) re: only pairwise comparisons of all framework residues (as defined by the IMTG definition)

Light: only pairwise comparisons of all framework residues (as defined by the IMTG definition)

As indicated by the above data, the humanized antibodies provided herein have significantly improved binding characteristics while having a favorable germinal ability index.

Example 2: CDR optimization

Many VH CDR and/or VL CDR variants were prepared. Binding affinity to recombinant human CXCR3 was measured using Biacore. Mutants with at least as strong binding as 53Hu37 are shown in Table 3.

Example 3: CXCR3-173 is a depleting antibody

Hamster anti-mouse CXCR3 monoclonal (clone CXCR3-173) was used as a surrogate antibody in preclinical experiments. CXCR3-173 has been previously described as a blocking antibody that does not deplete CD4+ T cells in vivo. (See Uppaluri et al., Transplantation 86:137-47 (2008)).

The following Fc variants of hamster CXCR3-173 mAb were prepared to test the effector function of the antibody: aglycosylated N297G variant of mouse IgG1 (CXCR3 mIgG1 agly); wild-type mouse IgG2a chimera (CXCR3 mIgG2a WT); Modified to eliminate capacity-depleting mouse IgG2a chimera (CXCR3 mIgG2a Dab); and wild-type mouse IgG3 (CXCR3mIgG3).

C57BL/6 mice (Jackson Laboratories, n=5 per group) aged 8 to 10 weeks were given a single 5 mg/kg intravenous dose of antibody and blood was harvested 24 hours later for flow cytometry using the following markers Cytometry analysis: CD4 and CD8 T cells by TCRab, CD4 and CD8; memory CD4 and CD8 T cells by TCRab, CD4, CD8, CD62L and CD44. Cells were quantified using Count Bright beads (Invitrogen) according to the manufacturer's protocol to determine the total number of cells per microliter of blood.

Gating results from total T lymphocytes and T lymphocyte subsets are shown in Figure 1. As shown, it was surprisingly demonstrated for the first time that hamster CXCR3-173 depletes CD4+ and CD8+ memory T cells when administered to mice.

Example 4: Effector functions of Fc variants of CXCR3-173

Mouse Fcγ receptor binding of the Fc-engineered form of CXCR3-173 was assessed using an antibody capture method using a Biacore 3000 instrument. Recombinant protein A/G (Pierce) was covalently immobilized onto a CM5 sensor chip using amine chemistry. The CXCR3-173 antibody was diluted to 5 μg/mL in HBS-EP buffer and injected into the protein A/G chip at a flow rate of 10 μL/min for 30 s. Recombinant mouse FcγRI (CD64), FcγRIIb (CD32), FcγRIII (CD16) and FcγIV (CD16-2) from R&D Systems were diluted 3-fold from 300 to 3.7 nM in HBS-EP buffer and prepared in duplicate as Inject the captured antibody at a flow rate of 30 μL/min. Surfaces were regenerated with glycine 2.0 (GE Healthcare). Binding responses were normalized to the amount of RU captured by protein A/G. The results are shown in Figures 2A-2D.

As shown, hamster CXCR3-173 modified to have the wild-type murine IgG2a isotype bound to all four recombinant mouse (rm) Fcγ receptors, although the dAB mutation significantly reduced this binding. CXCR3-173 modified to have the wild-type murine IgG3 isotype also bound to all four recombinant mouse Fcγ receptors. The original unmodified hamster CXCR3-173 bound rmFcγRIIb and rmFcγRIII better than the IgG2a isotype variant, whereas the non-glycosylated mIgG1 isotype variant did not bind any rmFcγR.

Example 5: Effector function in vitro

Humanized anti-CXCR3 mAbs and their Fc-engineered versions were studied in a series of ADCC assays. Fc-engineered versions of humanized anti-CXCR3 mAbs were prepared using standard methods. The defucosylated form was prepared by culturing cells expressing the humanized mAb in the presence of Kieff base.

Primary human NK cells or the NK9.2 cell line (Conkwest) overexpressing CD16 with a valine polymorphism were used as effector cells and CHO transfected cells overexpressing human CXCR3 (A isoform) were used as targets Cells were subjected to ADCC assay.

For assays using primary NK cells as effectors, NK cells were purified from leucopak of normal donors and cultured in IL-2 for 24 hours, then mixed with chromium-labeled overnight CHO- Human CXCR3 target cells were plated together. Cultures were incubated for 3 hours in a tissue culture incubator, then washed and lysed with 1% Tritron-X before reading the supernatants on a gamma counter.

For assays using NK9.2 cells as effectors, NK9.2 cells were expanded in IL-2 for 2 weeks following the manufacturer's recommendations. On the day of the assay, NK9.2 cells (70,000 cells) were labeled with Calcein AM (Invitrogen) and incubated with appropriately diluted antibody for 30 minutes to allow the antibody to bind to CXCR3 on the target cells. NK cells were plated at a 3:1 ratio of effector to target cells and the cultures were incubated for 1 hr in a tissue culture incubator. At the end of the culture period the cells were lysed with Triton X-100 and the plates were read using a M5 microplate reader (492nm excitation and 515nm emission).

Human IgGl (Sigma) was used as a negative control, and lysis of CD52-overexpressing CHO cells treated with alemtuzumab (monoclonal anti-CD52 antibody) served as a positive control for lysis. Signals are expressed in arbitrary fluorescence units (AFU). Percent cytotoxicity is expressed by (experimental lysis-spontaneous lysis)/(maximal lysis-spontaneous lysis) x 100%.

In the ADCC assay, humanized anti-CXCR3 mAb 53Hu37 with human IgGl Fc and Fc-engineered forms was tested. Fc-engineered forms M1 (S239D/D332E (EU notation), M2 (G236A/S267E/H268F/S324T/I332E (EU notation), "AEFTE") and M3 (S239D/H268F/S324T/I332E (EU notation method), "DFTE") contain amino acid changes that allow enhanced ADCC or CDC activity. A fourth Fc-engineered form was produced by Kiff base treatment of a cell line that produced the wild-type antibody.

Anti-human CXCR3 clones, clone 4 (CXCR3 CL4), clone 12 (CXCR3 CL12), clone 82 (CXCR3 CL82) and clone 135 (CXCR3 CL135) were also tested. In independent experiments, assays were performed with varying effector cells, effector:target (E:T) ratios, and antibody concentrations. Representative results are shown in Figure 3B and Figure 3C.

Figure 3A summarizes the effector functions of defucosylated forms of M1, M2, M3 and 53Hu37. Figure 3B shows the results of the assay using primary NK cells as effectors. Figure 3C shows the results of the assay using NK9.2 cells as effectors.

Example 6: Fcγ receptor binding

Human and mouse Fcγ receptor binding affinities of humanized anti-CXCR3 mAb 53Hu37 and Fc-engineered versions of 53Hu37 were assessed using a Biacore T200 instrument. Protein A from Sigma was immobilized on a CM5 series S chip using amine chemistry. Antibodies were injected into protein A chips, and various concentrations of recombinant human and mouse Fcγ receptors (R&D Systems) were injected into the captured antibodies. A wide concentration range of receptors was used spanning low and high affinity binders (1.2 nM to 5 μM). Each sample was injected in duplicate. Binding sensorgrams were fitted to a 1:1 kinetic binding model. The quantitative results are summarized in Figure 4.

As shown in Figure 4, the M1 and M3 Fc-engineered forms had improved affinity for both hFcγRIII and mFcγRIV, M2 had increased binding to hFcγRIIa, and Kiff base-treated 53Hu37 displayed hFcγRIII and mFcγRIIa compared to the Fc-engineered form. Moderate increase in mFcyRIV.

Embodiment 7:

Cynomolgus monkeys received a single intravenous infusion of 2 mg/kg body weight of 53Hu37, the Ml variant, or the Kiff base-treated form (n=8 per antibody treatment group) or vehicle control (n=6). Blood samples were collected before infusion and then 1, 3, 7, and 14 days after infusion, and analyzed by flow cytometry for total T cells and T cell subsets. For flow cytometry, erythrocytes were lysed and cells were stained with antibodies against the following markers to identify total and memory CD4 and CD8 T cells: CD3 (clone SP34-2), CD4 (clone OKT4), CD8a (clone RPA-T8), CD45RA (clone 5H9), CCR7 (clone G043H7) and CXCR3 (clone 1C6). Cells were quantified using Count Bright beads (Invitrogen) to determine the number of cells/microliter (cells/μL). Data are expressed as mean percentages of prebleed values for each cell subpopulation of the group over time. Histology of spleen samples obtained from subpopulations of each treatment group on day 14 post-infusion was studied by staining fixed sample sections for CXCR3 using anti-human CXCR3 clone 4 antibody and appropriate secondary antibodies. The results are shown in Figures 5A-5C. The pharmacokinetics of serum in blood samples were also determined using a peptide ELISA to measure circulating levels of administered antibodies.

As shown, 53Hu37, the M1 form of 53Hu37, and Kif-treated 53Hu37 reduced effector memory CD4 T cells, and 53Hu37, the M1 form of Hu37, and Kif-treated 53Hu37 reduced effector memory CD8 T cells. Furthermore, the M1 form of 53Hu37 and Kif-treated 53Hu37 significantly reduced CXCR3 staining in the spleen 14 days after a single intravenous infusion of the antibody.

Example 8: Biochemical Analysis

Thermal stability, stability against shear stress and virus inactivation, stability against freeze-thaw stress, stability against agitation stress, and pH stability of 53Hu37, the M1 form, and Kieff base-treated 53Hu37 were measured.

Differential Scanning Calorimetry (DSC)

Samples were analyzed with high-throughput VP-DSC (Microcal). Samples were diluted to approximately 0.5 mg/mL with the corresponding buffer and loaded onto 96-well plates. The scan parameters consisted of a start temperature of 25°C and an end temperature of 100°C. A scan rate of 200°C/h was used.

Turbidity

Sample absorbance at 340-360 nm was measured in 5 nm increments on a Spectramax Plus (Molecular Devices) and these values were averaged to generate the final turbidity measurement. Use a 96-well UV flat-bottom plate and 150-200 µL of material. Pre-read the plate before adding samples and apply path checks to the sample values. Based on "ATurbidimetric Method to Determine Visual Appearance of Protein Solutions" by Brigitte Eckhardt, Technology Applications, Vol. 48, No. 2, March-April 1994), comparing samples with values obtained from UV absorbance of turbidity standards A comparison is made to grade turbidity.

Size Exclusion Chromatography (SEC)

HP1100 or 1200 series systems are equipped with TSK SW _XL size exclusion columns coupled with SW _XL guard columns. Samples were run for 35 minutes using a mobile phase of 20 mM sodium phosphate, 500 mM NaCl, pH 6.0. A flow rate of 0.5 mL/min was used. A 50 μg injection was made and the UV signal was monitored at 280 nm.

Heat-Induced Relative Aggregation Propensity (TI-RAP)

0.2 mg/mL anti-human CXCR3 antibody (single antibody or antibody mixture) for 10 minutes to generate temperature-induced aggregation. After heat incubation, samples were centrifuged at 7000 xg for 2 minutes at 5°C to remove insoluble protein precipitates, and the supernatants were analyzed by cation exchange chromatography (CEX). The percentage of soluble monomer (and relative aggregation propensity) was calculated by normalizing the chromatographic peak area of the thermally stressed sample with the peak area of the control (5°C) sample.

Agitation-Induced Relative Aggregation Propensity (AI-RAP)

For agitation-induced relative aggregation propensity, antibodies containing 0.2 mg/mL final protein concentration were prepared in PBS buffer with 10 mM histidine and 9% sucrose buffer (containing 0 and 0.01% polysorbate 80) at 5°C. Solutions of human CXCR3 antibody were subjected to vigorous agitation stress at 5°C. The solution of anti-human CXCR3 antibody was agitated at maximum speed with a VX-2500 Multitube Vortexer (VWR, West Chester, PA) for a total of 24 hours.

Various time points throughout the study were analyzed by withdrawing small sample aliquots for CEX analysis. Sample aliquots were centrifuged at 7000 xg (at 5°C) for 2 minutes to remove insoluble protein precipitates, and the supernatants were analyzed by CEX. The percentage of soluble monomer (and relative aggregation propensity) was calculated by normalizing the chromatographic peak area of the agitated sample with the peak area of the control (0 hr) sample.

Cation Exchange Chromatography (CEX)

CEX analysis was performed on an Agilent 1290infinity HPLC system at 25°C using a ProPac WCX-10 analytical column (weak cation exchange, 4 x 250mm, Thermo Scientific). A 20 μg protein sample was loaded onto the column and analyzed at a flow rate of 0.8 mL/min. The column was equilibrated with buffer A (20 mM sodium acetate, 0.0025% sodium azide, pH 5.2) and buffer B (20 mM sodium acetate, 1 M sodium chloride, 0.0025% azide) with a linear gradient from 0 to 100%. NaCl, pH 5.2) to elute the protein for 40 minutes. The absorbance at 280nm was measured and the 280nm absorbance peak was integrated to determine the protein peak area.

experimental design

pH/temperature stress

A portion of material from each clone was dialyzed into 20 mM sodium phosphate at pH 5.0 and sodium phosphate at pH 7.0 using Slide-A-Lyzers (Thermo Scientific PN 66810). This material was diluted with the corresponding buffer and filtered to approximately 2 mg/mL using a Millex GV filter (Millipore PN SLGV033RB). Sodium phosphate samples at pH 5 and pH 7 were stored at 37°C for 3 and 5 weeks prior to testing.

freeze-thaw stress

The indicated anti-human CXCR3 antibodies were dialyzed into 20 mM sodium phosphate pH 6.0 using Slide-A-Lyzers (Thermo Scientific PN 66810). This material was diluted with the corresponding buffer and filtered to approximately 1 mg/mL using a Millex GV filter (Millipore PN SLGV033RB). Freeze-thaw stress samples were frozen at -80°C and thawed at room temperature a total of 5 times. After 1 and 3 freeze-thaw cycles, material was removed for testing by selection assay. A complete set of test assays was performed after the final freeze-thaw.

shear stress

Antibody samples were dialyzed into 20 mM sodium phosphate pH 6.0 using Slide-A-Lyzers (Thermo Scientific PN 66810). This material was diluted with the corresponding buffer and filtered to approximately 1 mg/mL using a Millex GV filter (Millipore PNSLGV033RB). The shear stress sample was repeatedly pipetted 50 times with a 200 μL pipette.

Simulated virus inactivation

Antibody samples were dialyzed into 20 mM sodium phosphate pH 6.0 using Slide-A-Lyzers (Thermo Scientific PN 66810). This material was diluted with the corresponding buffer and filtered to approximately 1 mg/mL using a Millex GV filter (Millipore PNSLGV033RB). Virus-inactivated samples were adjusted to pH 3.5 with 1N HCl and maintained at this pH for 100 min at room temperature. After the holding period, the samples were brought back to pH 7.2 using 1 N NaOH and held for 100 min. Samples were then adjusted to pH 6.0 using 1N HCl and tested.

result

Differential Scanning Calorimetry:

The M1 form (D/E mutant) was found to be slightly less stable than the Kif form, which in turn was slightly less stable than 53Hu37.

Shear stress:

The M1 form (D/E mutant) was found to be slightly less stable than the 53Hu37 and Kif forms, which were approximately equally stable.

Viral inactivation stress:

The M1 form (D/E mutant) was found to be more stable than the Kif form, both less stable than 53Hu37.

Freeze-thaw stress:

The M1 form (D/E mutant), Kif form and 53Hu37 were found to be roughly equivalent.

Agitation-Induced Relative Aggregation Tendency:

Approximately 80% of all forms of antibody precipitated within 2 hours of agitation, although at pH 5.5 the M1 form (D/E mutant) was relatively more stable than the other two forms.

Aggregate formation:

Under control (non-accelerated) conditions at pH 5.0, the M1 form (D/E mutant) and 53Hu37 were essentially stable over a period of 5 weeks, but the Kif form became milky; the same pattern was found under accelerated conditions . Under control (non-accelerated) conditions at pH 7.0, the three forms of the antibody were approximately the same over the 5 week period. Under accelerated conditions, the DE mutant was slightly more stable than 53Hu37, which in turn was more stable than the Kif form.

Example 9: Depleted but not non-depleted anti-CXCR3 is protective and therapeutic in a mouse model of vitiligo

mice. KRT14-Kitl*4XTG2Bjl (Krt14-Kitl*) mice were a gift from BJ Longley, University of Wisconsin. PMEL TCR transgenic mice were Thy1.1 ⁺ obtained from The Jackson Laboratory, stock number 005023, B6.Cg Thy1 ^a /CyTg(TcraTcrb)8Rest/J. All mice were maintained on a C57BL/6J background in a pathogen-free facility and procedures were performed according to the NIH Guide for the Care and Use of Laboratory Animals.

Antibody. ΔAB CXCR3 is an antibody that neutralizes CXCR3 activity (also referred to herein as IgG2a-dAB, dAB or ΔAB). WT mouse CXCR3 is an antibody to wild-type mouse IgG2a Fc with depletion activity.

Vitiligo induction and antibody therapy. Vitiligo was induced by adoptive transfer of premelanosome-specific (PMEL) CD8 ⁺ T cells as previously described. Harris et al., J. Invest. Dermatol. 132:1869-76 (2012). Briefly, PMEL CD8 ⁺ T cells were isolated from spleens of PMEL TCR transgenic mice (the Jackson Laboratory, stock number 005023) by negative selection on microbeads (Miltenyi Biotech) according to the manufacturer's instructions. Purified CD8 ⁺ T cells ( ¹ x 106 cells) were injected intravenously into sublethally irradiated (500 rads 1 day before transfer) Krt14-Kitl* hosts (theJackson Laboratory stock number 009687, 8-12 weeks old) . On the day of transfer, recipient mice also received an intraperitoneal injection of ¹ x 106 pfu rVV-hPMEL (N Restifo, NCI, NIH).

Treatment was performed by weekly intraperitoneal injections of 100 μg of antibody or isotype control three times during the observation period.

For prevention studies, mice were treated 2-6 weeks after vitiligo induction. This time period is important because it occurs after the virus used to induce disease is cleared, but before autoimmunity begins. Vitiligo scores were quantified using a score scale based on the degree of depigmentation at four easily visible locations, including the ears, nose, hind pads, and tail as previously described. Harris et al., J. Invest. Dermatol. 132:1869-76 (2012). Each location was examined and the degree of depigmentation was estimated as a percentage of the anatomical site; the left and right ears and the left and right hind pads were estimated together and thus evaluated as a single site. Scoring was determined as follows: no evidence of depigmentation (0%) scored 0, >0-10% = 1 point, >10%-25% = 2 points, >25%-75% = 3 points, >75%-< 100% = 4 points, 100% = 5 points. The "Vitiligo Score" is the sum of the scores from all four sites, with a maximum score of 20 points.

For repigmentation studies, mice with more than 75% of tail depigmentation were treated with WT mouse CXCR3-depleting Ab or isotype control at weeks 12–20 after vitiligo induction when the disease was normally stable. Pigment regeneration analysis was performed with ImageJ as previously described. Agarwal et al., J. Invest. Dermatol. 135:1080-8 (2015).

Effect of CXCR3 antibody on blood and spleen cell counts. Mice were infused with 100 μg of each antibody and assessed at 24 h for memory CD8 from blood (cells per microliter using Countbright beads) and spleen (total cell count using Via-Cell XR instrument from BectonCoulter) ⁺ T cell count. Because the administered antibody is essentially the same antibody (same CDR) as the antibody used for CXCR3 staining and could block binding of the stained antibody to CXCR3, use a CXCR3-independent measure by evaluating the number of memory CD8 ⁺ T cells to assess depletion (as defined by CD44 and CD62L staining), since more than 50% of these cells express CXCR3.

Flow Cytometry. Blood, tail, spleen, and skin draining lymph nodes were collected and analyzed by flow cytometry. Spleens were disrupted and erythrocytes were lysed using RBC Lysis Buffer (Sigma Aldrich). Blood samples were stained and processed with Biolegend Lysis/Fixation Buffer according to the manufacturer's protocol. To separate the dermis and epidermis, tail skin samples were incubated with 5 U mL ^-1 dispase II (Roche) for 1 hour at 37°C. Epidermis was removed and disrupted mechanically using a 70 μm cell strainer, and dermal samples were incubated with 1 mg mL ⁻¹ collagenase IV and 1–2 mg mL ⁻¹ DNAse I (Sigma-Aldrich) for 1 hour at 37°C. All cells were filtered through a 70 μm sieve prior to analysis. To identify PMELs and their phenotypes, samples were stained at 1:200 with the following antibodies in FACS buffer (1% BSA in PBS): CD45 AF700, CD8b PerCP-Cy5.5, Thy1.1 Pacific Blue, (Biolegend). Live cells were differentiated using Live/Dead Blue (Invitrogen) 1:1000 in FACS buffer. Data were collected and analyzed using a BD LSR II flow cytometer (BD Biosciences) and FlowJo software version 10 (Tree Star, Inc.).

Results: Effect of CXCR3 antibody on blood and spleen cell counts. Compared with untreated mice, in the blood and spleen, the total number of memory CD8 ⁺ T cells was significantly reduced in mice treated with WT mice or hamster CXCR3 antibodies, but not in mice treated with ΔAB CXCR3 Variety.

prevention. Mice were treated with hamster anti-mouse CXCR3 (depleted), WT mouse CXCR3 (superior depleted), ΔAB CXCR3 (neutralizing; non-depleted), or an isotype control antibody. Of these, CXCR3 in depleted antibody WT mice performed best in preventing clinical disease, as evidenced by a significantly reduced score 5 weeks after treatment (one-way ANOVA p=0.0066, Dunnett's post hoc test vs. isotype ns; Tukey's post hoc test for ΔAB Significant compared to WT and ΔAB compared to hamster). This observation is consistent with data indicating that WT mouse CXCR3 Ab has better depleting activity than hamster CXCR3 Ab. All Abs tested against vitiligo in mice resulted in less PMEL in the skin. Following treatment with any of the CXCR3 antibodies, PMEL numbers were significantly reduced in the epidermis (two-way ANOVA p<0.0001, Bonferroni post hoc test, compared to isotype controls) and tended to decrease in the dermis. However, despite their ability to reduce the number of PMELs in the skin, neutralizing antibodies were less effective than depleting antibodies, possibly due to the fact that low threshold numbers of PMELs are sufficient for full clinical disease. The effect of CXCR3-depleting antibodies on host T cells was assessed by measuring the number of host CD8 ⁺ T cells and total CD45 ⁺ cells in lymph nodes (LN), spleen, blood, epidermis and dermis. Bystander host CD8 ⁺ T cell numbers were significantly reduced in spleen and lymph nodes (LN) but not in skin by CXCR3 Ab treatment in hamster or WT mice (two-way ANOVA p<0.0001, Bonferroni post hoc test, vs. type control). However, the total number of CD45 ⁺ cells did not change after treatment with any CXCR3 Ab.

Pigment regeneration. Mouse WT CXCR3 Ab treatment significantly reversed clinical disease and reduced PMEL numbers in the epidermis. Although there was no change in total CD45 ⁺ cell numbers, treated mice had a slight reduction in host CD8 ⁺ T cell numbers. Taken together, these data indicate that a CXCR3-depleting Ab can reduce autoreactive T cell numbers and reverse disease with minimal impact on other compartments of the immune system.

The preceding examples are intended to illustrate and in no way limit the present disclosure. Other embodiments of the compounds and methods disclosed will be apparent to those skilled in the art from consideration of the specification and practice of the compounds and methods disclosed herein.

sequence listing

<110> Sanofi (SANOFI)

<120> Anti-human CXCR3 antibody for vitiligo treatment

<130> 597513: SA9-197PC

<150> 62/437,889

<151> 2016-12-22

<150> EP 17306770.3

<151> 2017-12-14

<160> 141

<170> PatentIn Version 3.5

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Met Val Leu Glu Val Ser Asp His Gln Val Leu Asn Asp Ala Glu Val

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Ala Ala Leu Leu Glu Asn Phe Ser Ser Ser Tyr Asp Tyr Gly Glu Asn

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Glu Ser Asp Ser Cys Cys Thr Ser Pro Pro Cys Pro Gln Asp Phe Ser

35 40 45

Leu Asn Phe Asp Arg Ala Phe Leu Pro Ala Leu Tyr Ser Leu Leu Phe

50 55 60

Leu Leu Gly Leu Leu Gly Asn Gly Ala Val Ala Ala Val Leu Leu Ser

65 70 75 80

Arg Arg Thr Ala Leu Ser Ser Thr Asp Thr Phe Leu Leu His Leu Ala

85 90 95

Val Ala Asp Thr Leu Leu Val Leu Thr Leu Pro Leu Trp Ala Val Asp

100 105 110

Ala Ala Val Gln Trp Val Phe Gly Ser Gly Leu Cys Lys Val Ala Gly

115 120 125

Ala Leu Phe Asn Ile Asn Phe Tyr Ala Gly Ala Leu Leu Leu Leu Ala Cys

130 135 140

Ile Ser Phe Asp Arg Tyr Leu Asn Ile Val His Ala Thr Gln Leu Tyr

145 150 155 160

Arg Arg Gly Pro Pro Ala Arg Val Thr Leu Thr Cys Leu Ala Val Trp

165 170 175

Gly Leu Cys Leu Leu Phe Ala Leu Pro Asp Phe Ile Phe Leu Ser Ala

180 185 190

His His Asp Glu Arg Leu Asn Ala Thr His Cys Gln Tyr Asn Phe Pro

195 200 205

Gln Val Gly Arg Thr Ala Leu Arg Val Leu Gln Leu Val Ala Gly Phe

210 215 220

Leu Leu Pro Leu Leu Val Met Ala Tyr Cys Tyr Ala His Ile Leu Ala

225 230 235 240

Val Leu Leu Val Ser Arg Gly Gln Arg Arg Leu Arg Ala Met Arg Leu

245 250 255

Val Val Val Val Val Val Ala Phe Ala Leu Cys Trp Thr Pro Tyr His

260 265 270

Leu Val Val Leu Val Asp Ile Leu Met Asp Leu Gly Ala Leu Ala Arg

275 280 285

Asn Cys Gly Arg Glu Ser Arg Val Asp Val Ala Lys Ser Val Thr Ser

290 295 300

Gly Leu Gly Tyr Met His Cys Cys Leu Asn Pro Leu Leu Tyr Ala Phe

305 310 315 320

Val Gly Val Lys Phe Arg Glu Arg Met Trp Met Leu Leu Leu Arg Leu

325 330 335

Gly Cys Pro Asn Gln Arg Gly Leu Gln Arg Gln Pro Ser Ser Ser Ser Arg

340 345 350

Arg Asp Ser Ser Trp Ser Glu Thr Ser Glu Ala Ser Tyr Ser Gly Leu

355 360 365

<210> 2

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<212> PRT

<213> Homo sapiens

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Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser

1 5 10 15

Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys Asp Lys Thr

20 25 30

His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser

35 40 45

Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg

50 55 60

Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro

65 70 75 80

Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala

85 90 95

Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val

100 105 110

Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr

115 120 125

Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr

130 135 140

Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu

145 150 155 160

Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys

165 170 175

Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser

180 185 190

Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp

195 200 205

Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser

210 215 220

Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala

225 230 235 240

Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly

245 250 255

<210> 3

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Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser

1 5 10 15

Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys Asp Lys Thr

20 25 30

His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Ala Gly Pro Ser

35 40 45

Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg

50 55 60

Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro

65 70 75 80

Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala

85 90 95

Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val

100 105 110

Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr

115 120 125

Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr

130 135 140

Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu

145 150 155 160

Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys

165 170 175

Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser

180 185 190

Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp

195 200 205

Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser

210 215 220

Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala

225 230 235 240

Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly

245 250 255

<210> 4

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Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser

1 5 10 15

Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys Asp Lys Thr

20 25 30

His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Asp

35 40 45

Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg

50 55 60

Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro

65 70 75 80

Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala

85 90 95

Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val

100 105 110

Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr

115 120 125

Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr

130 135 140

Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu

145 150 155 160

Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys

165 170 175

Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser

180 185 190

Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp

195 200 205

Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser

210 215 220

Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala

225 230 235 240

Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly

245 250 255

<210> 5

<211> 255

<212> PRT

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Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser

1 5 10 15

Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys Asp Lys Thr

20 25 30

His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser

35 40 45

Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg

50 55 60

Thr Pro Glu Val Thr Cys Val Val Val Asp Val Glu His Glu Asp Pro

65 70 75 80

Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala

85 90 95

Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val

100 105 110

Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr

115 120 125

Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr

130 135 140

Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu

145 150 155 160

Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys

165 170 175

Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser

180 185 190

Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp

195 200 205

Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser

210 215 220

Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala

225 230 235 240

Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly

245 250 255

<210> 6

<211> 255

<212> PRT

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Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser

1 5 10 15

Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys Asp Lys Thr

20 25 30

His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser

35 40 45

Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg

50 55 60

Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser Phe Glu Asp Pro

65 70 75 80

Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala

85 90 95

Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val

100 105 110

Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr

115 120 125

Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr

130 135 140

Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu

145 150 155 160

Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys

165 170 175

Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser

180 185 190

Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp

195 200 205

Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser

210 215 220

Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala

225 230 235 240

Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly

245 250 255

<210> 7

<211> 255

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<400> 7

Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser

1 5 10 15

Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys Asp Lys Thr

20 25 30

His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser

35 40 45

Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg

50 55 60

Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro

65 70 75 80

Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala

85 90 95

Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val

100 105 110

Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr

115 120 125

Lys Cys Lys Val Thr Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr

130 135 140

Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu

145 150 155 160

Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys

165 170 175

Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser

180 185 190

Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp

195 200 205

Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser

210 215 220

Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala

225 230 235 240

Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly

245 250 255

<210> 8

<211> 255

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 8

Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser

1 5 10 15

Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys Asp Lys Thr

20 25 30

His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser

35 40 45

Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg

50 55 60

Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro

65 70 75 80

Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala

85 90 95

Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val

100 105 110

Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr

115 120 125

Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Glu Glu Lys Thr

130 135 140

Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu

145 150 155 160

Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys

165 170 175

Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser

180 185 190

Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp

195 200 205

Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser

210 215 220

Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala

225 230 235 240

Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly

245 250 255

<210> 9

<211> 255

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 9

Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser

1 5 10 15

Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys Asp Lys Thr

20 25 30

His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Asp

35 40 45

Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg

50 55 60

Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro

65 70 75 80

Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala

85 90 95

Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val

100 105 110

Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr

115 120 125

Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Glu Glu Lys Thr

130 135 140

Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu

145 150 155 160

Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys

165 170 175

Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser

180 185 190

Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp

195 200 205

Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser

210 215 220

Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala

225 230 235 240

Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly

245 250 255

<210> 10

<211> 255

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 10

Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser

1 5 10 15

Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys Asp Lys Thr

20 25 30

His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Ala Gly Pro Ser

35 40 45

Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg

50 55 60

Thr Pro Glu Val Thr Cys Val Val Val Asp Val Glu Phe Glu Asp Pro

65 70 75 80

Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala

85 90 95

Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val

100 105 110

Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr

115 120 125

Lys Cys Lys Val Thr Asn Lys Ala Leu Pro Ala Pro Glu Glu Lys Thr

130 135 140

Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu

145 150 155 160

Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys

165 170 175

Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser

180 185 190

Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp

195 200 205

Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser

210 215 220

Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala

225 230 235 240

Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly

245 250 255

<210> 11

<211> 255

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 11

Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser

1 5 10 15

Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys Asp Lys Thr

20 25 30

His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Asp

35 40 45

Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile Ser Arg

50 55 60

Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser Phe Glu Asp Pro

65 70 75 80

Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala

85 90 95

Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val

100 105 110

Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr

115 120 125

Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Glu Glu Lys Thr

130 135 140

Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu

145 150 155 160

Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu Thr Cys

165 170 175

Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser

180 185 190

Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp

195 200 205

Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser

210 215 220

Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala

225 230 235 240

Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly

245 250 255

<210> 12

<211> 8

<212> PRT

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<220>

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<400> 12

Gly Phe Thr Phe Thr Ser Tyr Ala

1 5

<210> 13

<211> 8

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<220>

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<400> 13

Ile Ser His Gly Gly Thr Tyr Thr

1 5

<210> 14

<211> 16

<212> PRT

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<220>

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<400> 14

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

1 5 10 15

<210> 15

<211> 5

<212> PRT

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<400> 15

Ser Gly Val Asn Tyr

1 5

<210> 16

<211> 3

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<400> 16

Phe Thr Ser

1

<210> 17

<211> 9

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<400> 17

Gln Gln Phe Thr Ser Ser Ser Pro Tyr Thr

1 5

<210> 18

<211> 123

<212> PRT

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<220>

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<400> 18

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Val Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Ala Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 19

<211> 123

<212> PRT

<213> Artificial sequence

<220>

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<400> 19

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Met Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 20

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 20

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 21

<211> 106

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 21

Asp Ile Gln Leu Thr Gln Ser Pro Ser Phe Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Ser Gly Val Asn Tyr Leu

20 25 30

Tyr Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Trp Ile Tyr

35 40 45

Phe Thr Ser Thr Leu Ala Pro Gly Val Pro Ser Arg Phe Ser Gly Ser

50 55 60

Gly Ser Gly Asn Glu Tyr Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu

65 70 75 80

Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Phe Thr Ser Ser Pro Tyr Thr

85 90 95

Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys

100 105

<210> 22

<211> 106

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 22

Asp Ile Gln Leu Thr Gln Ser Pro Ser Phe Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Ser Gly Val Asn Tyr Leu

20 25 30

Tyr Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Trp Ile Tyr

35 40 45

Phe Thr Ser Thr Leu Ala Pro Gly Val Pro Ser Arg Phe Ser Gly Ser

50 55 60

Gly Ser Gly Asn Glu Tyr Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu

65 70 75 80

Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Phe Thr Ser Ser Pro Tyr Thr

85 90 95

Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys

100 105

<210> 23

<211> 106

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 23

Asp Ile Gln Leu Thr Gln Ser Pro Ser Phe Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Ser Gly Val Asn Tyr Leu

20 25 30

Tyr Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Trp Ile Tyr

35 40 45

Phe Thr Ser Thr Leu Ala Ser Gly Val Pro Ser Arg Phe Ser Gly Ser

50 55 60

Gly Ser Gly Asn Glu Tyr Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu

65 70 75 80

Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Phe Thr Ser Ser Pro Tyr Thr

85 90 95

Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys

100 105

<210> 24

<211> 106

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 24

Asp Ile Gln Leu Thr Gln Ser Pro Ser Phe Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Ser Gly Val Asn Tyr Leu

20 25 30

Tyr Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Trp Ile Tyr

35 40 45

Phe Thr Ser Thr Leu Ala Ser Gly Val Pro Ser Arg Phe Ser Gly Ser

50 55 60

Gly Ser Gly Asn Glu Tyr Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu

65 70 75 80

Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Phe Thr Ser Ser Pro Tyr Thr

85 90 95

Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys

100 105

<210> 25

<211> 452

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 25

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly

115 120 125

Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly

130 135 140

Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val

145 150 155 160

Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe

165 170 175

Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Ser Val Val

180 185 190

Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val

195 200 205

Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys

210 215 220

Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu

225 230 235 240

Leu Gly Gly Pro Asp Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr

245 250 255

Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val

260 265 270

Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val

275 280 285

Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser

290 295 300

Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu

305 310 315 320

Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala

325 330 335

Pro Glu Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro

340 345 350

Gln Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln

355 360 365

Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala

370 375 380

Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr

385 390 395 400

Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu

405 410 415

Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser

420 425 430

Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser

435 440 445

Leu Ser Pro Gly

450

<210> 26

<211> 452

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 26

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly

115 120 125

Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly

130 135 140

Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val

145 150 155 160

Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe

165 170 175

Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Ser Val Val

180 185 190

Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val

195 200 205

Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys

210 215 220

Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu

225 230 235 240

Leu Ala Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr

245 250 255

Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val

260 265 270

Glu Phe Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val

275 280 285

Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser

290 295 300

Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu

305 310 315 320

Asn Gly Lys Glu Tyr Lys Cys Lys Val Thr Asn Lys Ala Leu Pro Ala

325 330 335

Pro Glu Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro

340 345 350

Gln Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln

355 360 365

Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala

370 375 380

Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr

385 390 395 400

Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu

405 410 415

Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser

420 425 430

Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser

435 440 445

Leu Ser Pro Gly

450

<210> 27

<211> 452

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 27

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly

115 120 125

Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly

130 135 140

Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val

145 150 155 160

Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe

165 170 175

Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Ser Val Val

180 185 190

Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val

195 200 205

Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys

210 215 220

Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu

225 230 235 240

Leu Gly Gly Pro Asp Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr

245 250 255

Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val

260 265 270

Ser Phe Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val

275 280 285

Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser

290 295 300

Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu

305 310 315 320

Asn Gly Lys Glu Tyr Lys Cys Lys Val Thr Asn Lys Ala Leu Pro Ala

325 330 335

Pro Glu Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro

340 345 350

Gln Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln

355 360 365

Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala

370 375 380

Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr

385 390 395 400

Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu

405 410 415

Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser

420 425 430

Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser

435 440 445

Leu Ser Pro Gly

450

<210> 28

<211> 452

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 28

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Met Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly

115 120 125

Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly

130 135 140

Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val

145 150 155 160

Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe

165 170 175

Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Ser Val Val

180 185 190

Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val

195 200 205

Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys

210 215 220

Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu

225 230 235 240

Leu Gly Gly Pro Asp Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr

245 250 255

Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val

260 265 270

Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val

275 280 285

Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser

290 295 300

Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu

305 310 315 320

Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala

325 330 335

Pro Glu Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro

340 345 350

Gln Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln

355 360 365

Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala

370 375 380

Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr

385 390 395 400

Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu

405 410 415

Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser

420 425 430

Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser

435 440 445

Leu Ser Pro Gly

450

<210> 29

<211> 452

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 29

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Met Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly

115 120 125

Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly

130 135 140

Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val

145 150 155 160

Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe

165 170 175

Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Ser Val Val

180 185 190

Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val

195 200 205

Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys

210 215 220

Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu

225 230 235 240

Leu Ala Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr

245 250 255

Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val

260 265 270

Glu Phe Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val

275 280 285

Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser

290 295 300

Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu

305 310 315 320

Asn Gly Lys Glu Tyr Lys Cys Lys Val Thr Asn Lys Ala Leu Pro Ala

325 330 335

Pro Glu Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro

340 345 350

Gln Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln

355 360 365

Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala

370 375 380

Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr

385 390 395 400

Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu

405 410 415

Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser

420 425 430

Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser

435 440 445

Leu Ser Pro Gly

450

<210> 30

<211> 452

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 30

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Met Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly

115 120 125

Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly

130 135 140

Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val

145 150 155 160

Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe

165 170 175

Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Ser Val Val

180 185 190

Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val

195 200 205

Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys

210 215 220

Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu

225 230 235 240

Leu Gly Gly Pro Asp Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr

245 250 255

Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val

260 265 270

Ser Phe Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val

275 280 285

Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser

290 295 300

Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu

305 310 315 320

Asn Gly Lys Glu Tyr Lys Cys Lys Val Thr Asn Lys Ala Leu Pro Ala

325 330 335

Pro Glu Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro

340 345 350

Gln Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln

355 360 365

Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala

370 375 380

Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr

385 390 395 400

Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu

405 410 415

Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser

420 425 430

Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser

435 440 445

Leu Ser Pro Gly

450

<210> 31

<211> 452

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 31

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Val Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Ala Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly

115 120 125

Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly

130 135 140

Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val

145 150 155 160

Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe

165 170 175

Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Ser Val Val

180 185 190

Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val

195 200 205

Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys

210 215 220

Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu

225 230 235 240

Leu Gly Gly Pro Asp Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr

245 250 255

Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val

260 265 270

Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val

275 280 285

Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser

290 295 300

Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu

305 310 315 320

Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala

325 330 335

Pro Glu Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro

340 345 350

Gln Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln

355 360 365

Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala

370 375 380

Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr

385 390 395 400

Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu

405 410 415

Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser

420 425 430

Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser

435 440 445

Leu Ser Pro Gly

450

<210> 32

<211> 452

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 32

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Val Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Ala Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly

115 120 125

Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly

130 135 140

Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val

145 150 155 160

Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe

165 170 175

Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Ser Val Val

180 185 190

Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val

195 200 205

Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys

210 215 220

Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu

225 230 235 240

Leu Ala Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr

245 250 255

Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val

260 265 270

Glu Phe Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val

275 280 285

Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser

290 295 300

Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu

305 310 315 320

Asn Gly Lys Glu Tyr Lys Cys Lys Val Thr Asn Lys Ala Leu Pro Ala

325 330 335

Pro Glu Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro

340 345 350

Gln Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln

355 360 365

Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala

370 375 380

Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr

385 390 395 400

Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu

405 410 415

Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser

420 425 430

Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser

435 440 445

Leu Ser Pro Gly

450

<210> 33

<211> 452

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 33

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Val Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Ala Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly

115 120 125

Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly

130 135 140

Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val

145 150 155 160

Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe

165 170 175

Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Ser Val Val

180 185 190

Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val

195 200 205

Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys

210 215 220

Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu

225 230 235 240

Leu Gly Gly Pro Asp Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr

245 250 255

Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val

260 265 270

Ser Phe Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val

275 280 285

Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser

290 295 300

Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu

305 310 315 320

Asn Gly Lys Glu Tyr Lys Cys Lys Val Thr Asn Lys Ala Leu Pro Ala

325 330 335

Pro Glu Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro

340 345 350

Gln Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln

355 360 365

Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala

370 375 380

Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr

385 390 395 400

Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu

405 410 415

Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser

420 425 430

Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser

435 440 445

Leu Ser Pro Gly

450

<210> 34

<211> 8

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 34

Gly Phe Thr Phe Arg Ser Tyr Ala

1 5

<210> 35

<211> 8

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 35

Ile Ser His Lys Gly Thr Tyr Thr

1 5

<210> 36

<211> 8

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 36

Ile Ser His Lys Gly Lys Tyr Thr

1 5

<210> 37

<211> 8

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 37

Ile Ser His Arg Gly Thr Tyr Thr

1 5

<210> 38

<211> 8

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 38

Ile Ser His Arg Gly Arg Tyr Thr

1 5

<210> 39

<211> 8

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 39

Ile Ser Arg Gly Gly Thr Tyr Thr

1 5

<210> 40

<211> 8

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 40

Ile Ser Ser Gly Gly Thr Tyr Thr

1 5

<210> 41

<211> 8

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 41

Ile Ser His Gly Gly Lys Tyr Thr

1 5

<210> 42

<211> 8

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 42

Ile Ser His Gly Gly Arg Tyr Thr

1 5

<210> 43

<211> 8

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 43

Ile Ser His Gly Gly Trp Tyr Thr

1 5

<210> 44

<211> 8

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 44

Ile Ser His Gly Gly Tyr Tyr Thr

1 5

<210> 45

<211> 16

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 45

Ala Arg His Pro Ile Tyr Ser Gly Phe Tyr Gln Gly Tyr Phe Asp Tyr

1 5 10 15

<210> 46

<211> 16

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 46

Ala Arg His Pro Ile Tyr Ser Gly Trp Tyr Gln Gly Tyr Phe Asp Tyr

1 5 10 15

<210> 47

<211> 16

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 47

Ala Arg His Pro Ile Tyr Ser Gly Gln Tyr Gln Gly Tyr Phe Asp Tyr

1 5 10 15

<210> 48

<211> 16

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 48

Ala Arg His Pro Ile Tyr Ala Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

1 5 10 15

<210> 49

<211> 16

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 49

Ala Arg His Pro Ile Tyr Cys Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

1 5 10 15

<210> 50

<211> 16

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 50

Ala Arg His Pro Ile Tyr Asp Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

1 5 10 15

<210> 51

<211> 16

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 51

Ala Arg His Pro Ile Tyr Glu Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

1 5 10 15

<210> 52

<211> 16

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 52

Ala Arg His Pro Ile Tyr Phe Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

1 5 10 15

<210> 53

<211> 16

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 53

Ala Arg His Pro Ile Tyr Gly Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

1 5 10 15

<210> 54

<211> 16

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 54

Ala Arg His Pro Ile Tyr His Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

1 5 10 15

<210> 55

<211> 16

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 55

Ala Arg His Pro Ile Tyr Ile Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

1 5 10 15

<210> 56

<211> 16

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 56

Ala Arg His Pro Ile Tyr Lys Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

1 5 10 15

<210> 57

<211> 16

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 57

Ala Arg His Pro Ile Tyr Leu Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

1 5 10 15

<210> 58

<211> 16

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 58

Ala Arg His Pro Ile Tyr Met Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

1 5 10 15

<210> 59

<211> 16

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 59

Ala Arg His Pro Ile Tyr Asn Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

1 5 10 15

<210> 60

<211> 16

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 60

Ala Arg His Pro Ile Tyr Pro Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

1 5 10 15

<210> 61

<211> 16

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 61

Ala Arg His Pro Ile Tyr Gln Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

1 5 10 15

<210> 62

<211> 16

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 62

Ala Arg His Pro Ile Tyr Arg Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

1 5 10 15

<210> 63

<211> 16

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 63

Ala Arg His Pro Ile Tyr Thr Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

1 5 10 15

<210> 64

<211> 16

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 64

Ala Arg His Pro Ile Tyr Val Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

1 5 10 15

<210> 65

<211> 16

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 65

Ala Arg His Pro Ile Tyr Trp Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

1 5 10 15

<210> 66

<211> 16

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 66

Ala Arg His Pro Ile Tyr Tyr Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

1 5 10 15

<210> 67

<211> 8

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 67

Ile Ser His Gly Gly Arg Tyr Thr

1 5

<210> 68

<211> 16

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 68

Ala Arg His Pro Ile His Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

1 5 10 15

<210> 69

<211> 5

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 69

Ser Gly Val Ile Tyr

1 5

<210> 70

<211> 5

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 70

Ser Gly Val Lys Tyr

1 5

<210> 71

<211> 5

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 71

Ser Gly Val Arg Tyr

1 5

<210> 72

<211> 5

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 72

Ser Gly Val Ser Tyr

1 5

<210> 73

<211> 5

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 73

Ser Gly Val Trp Tyr

1 5

<210> 74

<211> 5

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 74

Ser Gly Val Tyr Tyr

1 5

<210> 75

<211> 9

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 75

Gln Gln Phe Thr Arg Ser Pro Tyr Thr

1 5

<210> 76

<211> 9

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 76

Gln Gln Phe Thr Ser Lys Pro Tyr Thr

1 5

<210> 77

<211> 9

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 77

Gln Gln Phe Lys Ser Ser Pro Tyr Thr

1 5

<210> 78

<211> 9

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 78

Gln Gln Phe Arg Ser Ser Pro Tyr Thr

1 5

<210> 79

<211> 9

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 79

Gln Gln Phe Tyr Ser Ser Pro Tyr Thr

1 5

<210> 80

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 80

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Lys Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 81

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 81

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Lys Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Phe Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 82

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 82

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Lys Gly Lys Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Phe Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 83

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 83

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Arg Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 84

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 84

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Arg Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Phe Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 85

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 85

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Arg Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Trp Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 86

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 86

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Arg Gly Arg Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 87

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 87

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Arg Gly Arg Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Phe Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 88

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 88

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser Arg Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 89

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 89

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser Arg Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Gln Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 90

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 90

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser Ser Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 91

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 91

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Phe Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 92

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 92

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Trp Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 93

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 93

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ala Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 94

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 94

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Cys Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 95

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 95

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Asp Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 96

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 96

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Glu Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 97

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 97

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Phe Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 98

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 98

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Gly Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 99

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 99

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr His Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 100

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 100

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ile Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 101

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 101

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Lys Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 102

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 102

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Leu Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 103

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 103

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Met Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 104

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 104

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Asn Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 105

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 105

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Pro Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 106

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 106

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Gln Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 107

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 107

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Arg Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 108

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 108

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Thr Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 109

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 109

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Val Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 110

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 110

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Trp Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 111

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 111

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Tyr Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 112

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 112

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Arg Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 113

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 113

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Lys Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 114

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 114

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Lys Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Trp Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 115

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 115

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Arg Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 116

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 116

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Arg Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Phe Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 117

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 117

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Trp Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 118

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 118

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Tyr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 119

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 119

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Lys Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 120

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 120

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile His Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 121

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 121

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Arg Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 122

<211> 123

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 122

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Trp Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser

115 120

<210> 123

<211> 106

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 123

Asp Ile Gln Leu Thr Gln Ser Pro Ser Phe Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Ser Gly Val Ile Tyr Leu

20 25 30

Tyr Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Trp Ile Tyr

35 40 45

Phe Thr Ser Thr Leu Ala Ser Gly Val Pro Ser Arg Phe Ser Gly Ser

50 55 60

Gly Ser Gly Asn Glu Tyr Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu

65 70 75 80

Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Phe Thr Ser Ser Pro Tyr Thr

85 90 95

Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys

100 105

<210> 124

<211> 106

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 124

Asp Ile Gln Leu Thr Gln Ser Pro Ser Phe Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Ser Gly Val Lys Tyr Leu

20 25 30

Tyr Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Trp Ile Tyr

35 40 45

Phe Thr Ser Thr Leu Ala Ser Gly Val Pro Ser Arg Phe Ser Gly Ser

50 55 60

Gly Ser Gly Asn Glu Tyr Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu

65 70 75 80

Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Phe Thr Ser Ser Pro Tyr Thr

85 90 95

Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys

100 105

<210> 125

<211> 106

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 125

Asp Ile Gln Leu Thr Gln Ser Pro Ser Phe Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Ser Gly Val Arg Tyr Leu

20 25 30

Tyr Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Trp Ile Tyr

35 40 45

Phe Thr Ser Thr Leu Ala Ser Gly Val Pro Ser Arg Phe Ser Gly Ser

50 55 60

Gly Ser Gly Asn Glu Tyr Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu

65 70 75 80

Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Phe Thr Ser Ser Pro Tyr Thr

85 90 95

Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys

100 105

<210> 126

<211> 106

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 126

Asp Ile Gln Leu Thr Gln Ser Pro Ser Phe Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Ser Gly Val Arg Tyr Leu

20 25 30

Tyr Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Trp Ile Tyr

35 40 45

Phe Thr Ser Thr Leu Ala Ser Gly Val Pro Ser Arg Phe Ser Gly Ser

50 55 60

Gly Ser Gly Asn Glu Tyr Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu

65 70 75 80

Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Phe Thr Arg Ser Pro Tyr Thr

85 90 95

Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys

100 105

<210> 127

<211> 106

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 127

Asp Ile Gln Leu Thr Gln Ser Pro Ser Phe Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Ser Gly Val Ser Tyr Leu

20 25 30

Tyr Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Trp Ile Tyr

35 40 45

Phe Thr Ser Thr Leu Ala Ser Gly Val Pro Ser Arg Phe Ser Gly Ser

50 55 60

Gly Ser Gly Asn Glu Tyr Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu

65 70 75 80

Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Phe Thr Ser Ser Pro Tyr Thr

85 90 95

Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys

100 105

<210> 128

<211> 106

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 128

Asp Ile Gln Leu Thr Gln Ser Pro Ser Phe Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Ser Gly Val Trp Tyr Leu

20 25 30

Tyr Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Trp Ile Tyr

35 40 45

Phe Thr Ser Thr Leu Ala Ser Gly Val Pro Ser Arg Phe Ser Gly Ser

50 55 60

Gly Ser Gly Asn Glu Tyr Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu

65 70 75 80

Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Phe Thr Ser Ser Pro Tyr Thr

85 90 95

Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys

100 105

<210> 129

<211> 106

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 129

Asp Ile Gln Leu Thr Gln Ser Pro Ser Phe Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Ser Gly Val Tyr Tyr Leu

20 25 30

Tyr Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Trp Ile Tyr

35 40 45

Phe Thr Ser Thr Leu Ala Ser Gly Val Pro Ser Arg Phe Ser Gly Ser

50 55 60

Gly Ser Gly Asn Glu Tyr Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu

65 70 75 80

Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Phe Thr Ser Ser Pro Tyr Thr

85 90 95

Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys

100 105

<210> 130

<211> 106

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 130

Asp Ile Gln Leu Thr Gln Ser Pro Ser Phe Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Ser Gly Val Asn Tyr Leu

20 25 30

Tyr Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Trp Ile Tyr

35 40 45

Phe Thr Ser Thr Leu Ala Ser Gly Val Pro Ser Arg Phe Ser Gly Ser

50 55 60

Gly Ser Gly Asn Glu Tyr Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu

65 70 75 80

Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Phe Thr Arg Ser Pro Tyr Thr

85 90 95

Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys

100 105

<210> 131

<211> 106

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 131

Asp Ile Gln Leu Thr Gln Ser Pro Ser Phe Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Ser Gly Val Asn Tyr Leu

20 25 30

Tyr Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Trp Ile Tyr

35 40 45

Phe Thr Ser Thr Leu Ala Ser Gly Val Pro Ser Arg Phe Ser Gly Ser

50 55 60

Gly Ser Gly Asn Glu Tyr Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu

65 70 75 80

Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Phe Thr Ser Lys Pro Tyr Thr

85 90 95

Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys

100 105

<210> 132

<211> 106

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 132

Asp Ile Gln Leu Thr Gln Ser Pro Ser Phe Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Ser Gly Val Asn Tyr Leu

20 25 30

Tyr Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Trp Ile Tyr

35 40 45

Phe Thr Ser Thr Leu Ala Ser Gly Val Pro Ser Arg Phe Ser Gly Ser

50 55 60

Gly Ser Gly Asn Glu Tyr Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu

65 70 75 80

Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Phe Lys Ser Ser Pro Tyr Thr

85 90 95

Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys

100 105

<210> 133

<211> 106

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 133

Asp Ile Gln Leu Thr Gln Ser Pro Ser Phe Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Ser Gly Val Asn Tyr Leu

20 25 30

Tyr Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Trp Ile Tyr

35 40 45

Phe Thr Ser Thr Leu Ala Ser Gly Val Pro Ser Arg Phe Ser Gly Ser

50 55 60

Gly Ser Gly Asn Glu Tyr Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu

65 70 75 80

Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Phe Arg Ser Ser Pro Tyr Thr

85 90 95

Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys

100 105

<210> 134

<211> 106

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 134

Asp Ile Gln Leu Thr Gln Ser Pro Ser Phe Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Ser Gly Val Asn Tyr Leu

20 25 30

Tyr Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Trp Ile Tyr

35 40 45

Phe Thr Ser Thr Leu Ala Ser Gly Val Pro Ser Arg Phe Ser Gly Ser

50 55 60

Gly Ser Gly Asn Glu Tyr Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu

65 70 75 80

Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Phe Tyr Ser Ser Pro Tyr Thr

85 90 95

Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys

100 105

<210> 135

<211> 213

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 135

Asp Ile Gln Leu Thr Gln Ser Pro Ser Phe Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Ser Gly Val Asn Tyr Leu

20 25 30

Tyr Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Trp Ile Tyr

35 40 45

Phe Thr Ser Thr Leu Ala Ser Gly Val Pro Ser Arg Phe Ser Gly Ser

50 55 60

Gly Ser Gly Asn Glu Tyr Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu

65 70 75 80

Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Phe Thr Ser Ser Pro Tyr Thr

85 90 95

Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys Arg Thr Val Ala Ala Pro

100 105 110

Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly Thr

115 120 125

Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala Lys

130 135 140

Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln Glu

145 150 155 160

Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser Ser

165 170 175

Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr Ala

180 185 190

Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser Phe

195 200 205

Asn Arg Gly Glu Cys

210

<210> 136

<211> 213

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 136

Asp Ile Gln Leu Thr Gln Ser Pro Ser Phe Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Ser Gly Val Asn Tyr Leu

20 25 30

Tyr Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Trp Ile Tyr

35 40 45

Phe Thr Ser Thr Leu Ala Ser Gly Val Pro Ser Arg Phe Ser Gly Ser

50 55 60

Gly Ser Gly Asn Glu Tyr Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu

65 70 75 80

Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Phe Thr Ser Ser Pro Tyr Thr

85 90 95

Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys Arg Thr Val Ala Ala Pro

100 105 110

Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly Thr

115 120 125

Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala Lys

130 135 140

Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln Glu

145 150 155 160

Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser Ser

165 170 175

Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr Ala

180 185 190

Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser Phe

195 200 205

Asn Arg Gly Glu Cys

210

<210> 137

<211> 213

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 137

Asp Ile Gln Leu Thr Gln Ser Pro Ser Phe Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Ser Gly Val Asn Tyr Leu

20 25 30

Tyr Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Trp Ile Tyr

35 40 45

Phe Thr Ser Thr Leu Ala Pro Gly Val Pro Ser Arg Phe Ser Gly Ser

50 55 60

Gly Ser Gly Asn Glu Tyr Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu

65 70 75 80

Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Phe Thr Ser Ser Pro Tyr Thr

85 90 95

Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys Arg Thr Val Ala Ala Pro

100 105 110

Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly Thr

115 120 125

Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala Lys

130 135 140

Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln Glu

145 150 155 160

Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser Ser

165 170 175

Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr Ala

180 185 190

Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser Phe

195 200 205

Asn Arg Gly Glu Cys

210

<210> 138

<211> 213

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 138

Asp Ile Gln Leu Thr Gln Ser Pro Ser Phe Leu Ser Ala Ser Val Gly

1 5 10 15

Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Ser Gly Val Asn Tyr Leu

20 25 30

Tyr Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Trp Ile Tyr

35 40 45

Phe Thr Ser Thr Leu Ala Pro Gly Val Pro Ser Arg Phe Ser Gly Ser

50 55 60

Gly Ser Gly Asn Glu Tyr Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu

65 70 75 80

Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Phe Thr Ser Ser Pro Tyr Thr

85 90 95

Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys Arg Thr Val Ala Ala Pro

100 105 110

Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly Thr

115 120 125

Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala Lys

130 135 140

Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser Gln Glu

145 150 155 160

Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser Ser

165 170 175

Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr Ala

180 185 190

Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys Ser Phe

195 200 205

Asn Arg Gly Glu Cys

210

<210> 139

<211> 452

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 139

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly

115 120 125

Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly

130 135 140

Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val

145 150 155 160

Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe

165 170 175

Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Ser Val Val

180 185 190

Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val

195 200 205

Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys

210 215 220

Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu

225 230 235 240

Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr

245 250 255

Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val

260 265 270

Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val

275 280 285

Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser

290 295 300

Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu

305 310 315 320

Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala

325 330 335

Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro

340 345 350

Gln Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln

355 360 365

Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala

370 375 380

Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr

385 390 395 400

Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu

405 410 415

Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser

420 425 430

Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser

435 440 445

Leu Ser Pro Gly

450

<210> 140

<211> 452

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 140

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Val Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Ala Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly

115 120 125

Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly

130 135 140

Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val

145 150 155 160

Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe

165 170 175

Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Ser Val Val

180 185 190

Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val

195 200 205

Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys

210 215 220

Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu

225 230 235 240

Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr

245 250 255

Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val

260 265 270

Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val

275 280 285

Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser

290 295 300

Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu

305 310 315 320

Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala

325 330 335

Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro

340 345 350

Gln Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln

355 360 365

Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala

370 375 380

Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr

385 390 395 400

Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu

405 410 415

Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser

420 425 430

Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser

435 440 445

Leu Ser Pro Gly

450

<210> 141

<211> 452

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic peptides

<400> 141

Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly

1 5 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Thr Ser Tyr

20 25 30

Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val

35 40 45

Ala Thr Ile Ser His Gly Gly Thr Tyr Thr Tyr Tyr Pro Asp Ser Val

50 55 60

Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Thr Leu Tyr

65 70 75 80

Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Met Tyr Tyr Cys

85 90 95

Ala Arg His Pro Ile Tyr Ser Gly Asn Tyr Gln Gly Tyr Phe Asp Tyr

100 105 110

Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly

115 120 125

Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly

130 135 140

Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val

145 150 155 160

Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe

165 170 175

Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Ser Val Val

180 185 190

Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val

195 200 205

Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys

210 215 220

Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu

225 230 235 240

Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr

245 250 255

Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val

260 265 270

Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val

275 280 285

Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser

290 295 300

Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu

305 310 315 320

Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala

325 330 335

Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro

340 345 350

Gln Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln

355 360 365

Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala

370 375 380

Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr

385 390 395 400

Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu

405 410 415

Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser

420 425 430

Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser

435 440 445

Leu Ser Pro Gly

450

Claims (32)

1. Humanized anti-human C-X-C motif CC chemokine receptor 3 used in a kind of method in treatment leucoderma (CXCR3) antibody or its pharmaceutical composition, wherein the Humanized anti-human CXCR3 antibody includes with heavy chain variable region (VH) Heavy chain (HC) and the light chain (LC) with light chain variable region (VL), wherein

A) VH and VL include be respectively selected from SEQ ID NO:24/20,22/18,80/130,81/24,82/130,82/24, 83/126、83/130、83/24、84/126、84/24、85/24、86/126、87/126、87/133、87/24、88/24、89/ 24、90/126、91/126、91/130、92/130、92/24、93/126、94/126、95/126、96/126、97/126、98/ 126、99/126、100/126、101/126、102/126、103/126、104/126、105/126、106/126、107/126、 108/126、109/126、110/126、111/126、121/24、113/130、113/24、114/130、115/126、115/ 130、115/24、116/126、116/130、116/24、117/126、118/126、20/123、20/124、20/125、20/ 126、20/127、20/128、20/129、20/130、20/131、20/132、20/133、20/134、119/126、122/126、 With the amino acid sequence of 120/130 sequence pair, and wherein

B) heavy chain include the area human IgG1 Fc, the area the human IgG1 Fc have selected from SEQ ID NO:2,3,4,5,6,7,8, 9,10 and 11 amino acid sequence.

2. the Humanized anti-human CXCR3 antibody of claim 1, wherein the VH includes the amino acid sequence of SEQ ID NO:20, And the VL includes the amino acid sequence of SEQ ID NO:24.

3. the Humanized anti-human CXCR3 antibody of claim 2, wherein the area the human IgG1 Fc includes the amino of SEQ ID NO:2 Acid sequence.

4. the Humanized anti-human CXCR3 antibody of claim 2, wherein the area the human IgG1 Fc includes the amino of SEQ ID NO:9 Acid sequence.

5. the Humanized anti-human CXCR3 antibody of claim 2, wherein the area the human IgG1 Fc includes the ammonia of SEQ ID NO:10 Base acid sequence.

6. the Humanized anti-human CXCR3 antibody of claim 2, wherein the area the human IgG1 Fc includes the ammonia of SEQ ID NO:11 Base acid sequence.

7. the Humanized anti-human CXCR3 antibody of claim 1, wherein the VH includes the amino acid sequence of SEQ ID NO:18, And the VL includes the amino acid sequence of SEQ ID NO:22.

8. the Humanized anti-human CXCR3 antibody of claim 7, wherein the area the human IgG1 Fc includes the amino of SEQ ID NO:2 Acid sequence.

9. the Humanized anti-human CXCR3 antibody of claim 8, wherein the area the human IgG1 Fc includes the amino of SEQ ID NO:9 Acid sequence.

10. the Humanized anti-human CXCR3 antibody of claim 8, wherein the area the human IgG1 Fc includes the ammonia of SEQ ID NO:10 Base acid sequence.

11. the Humanized anti-human CXCR3 antibody of claim 8, wherein the area the human IgG1 Fc includes the ammonia of SEQ ID NO:11 Base acid sequence.

12. the Humanized anti-human CXCR3 antibody or its medicament preparation of claim 35, wherein

A) HC includes the amino acid sequence of SEQ ID NO:25, and the LC includes the amino acid of SEQ ID NO:135 Sequence,

B) HC includes the amino acid sequence of SEQ ID NO:26, and the LC includes the amino acid of SEQ ID NO:135 Sequence,

C) HC includes the amino acid sequence of SEQ ID NO:27, and the LC includes the amino acid of SEQ ID NO:135 Sequence,

D) HC includes the amino acid sequence of SEQ ID NO:139, and the LC includes the amino acid of SEQ ID NO:135 Sequence,

E) HC includes the amino acid sequence of SEQ ID NO:31, and the LC includes the amino acid of SEQ ID NO:137 Sequence,

F) HC includes the amino acid sequence of SEQ ID NO:32, and the LC includes the amino acid of SEQ ID NO:137 Sequence,

G) HC includes the amino acid sequence of SEQ ID NO:32, and the LC includes the amino acid of SEQ ID NO:137 Sequence,

H) HC includes the amino acid sequence of SEQ ID NO:140, and the HC includes the amino acid of SEQ ID NO:137 Sequence.

13. the Humanized anti-human CXCR3 antibody or its medicament preparation of any one of claim 1-12, wherein the IgG1 The area Fc has reduced fucose content.

14. the Humanized anti-human CXCR3 antibody or its medicament preparation of any one of claim 1-12, wherein the antibody is It is generated in following host cell, the host cell is cultivated in the presence of glycosylation inhibitor.

15. the Humanized anti-human CXCR3 antibody or its medicament preparation of claim 14, wherein the glycosylation inhibitor is base Husband's alkali.

16. a kind of method for treating leucoderma comprising comprising the heavy chain (HC) with heavy chain variable region (VH) and will have light The Humanized anti-human CXCR3 antibody of the light chain (LC) of chain variable region (VL) is given to subject in need thereof, wherein

A) VH and VL include be respectively selected from SEQ ID NO:24/20,22/18,80/130,81/24,82/130,82/24, 83/126、83/130、83/24、84/126、84/24、85/24、86/126、87/126、87/133、87/24、88/24、89/ 24、90/126、91/126、91/130、92/130、92/24、93/126、94/126、95/126、96/126、97/126、98/ 126、99/126、100/126、101/126、102/126、103/126、104/126、105/126、106/126、107/126、 108/126、109/126、110/126、111/126、121/24、113/130、113/24、114/130、115/126、115/ 130、115/24、116/126、116/130、116/24、117/126、118/126、20/123、20/124、20/125、20/ 126、20/127、20/128、20/129、20/130、20/131、20/132、20/133、20/134、119/126、122/126、 With the amino acid sequence of 120/130 sequence pair, and wherein

B) heavy chain includes the area human IgG1 Fc selected from SEQ ID NO:2,3,4,5,6,7,8,9,10 and 11.

17. the method for claim 16, wherein the VH includes the amino acid sequence of SEQ ID NO:20, and the VL includes The amino acid sequence of SEQ ID NO:24.

18. the Humanized anti-human CXCR3 antibody of claim 17, wherein the area the human IgG1 Fc includes the ammonia of SEQ ID NO:2 Base acid sequence.

19. the Humanized anti-human CXCR3 antibody of claim 17, wherein the area the human IgG1 Fc includes the ammonia of SEQ ID NO:9 Base acid sequence.

20. the Humanized anti-human CXCR3 antibody of claim 17, wherein the area the human IgG1 Fc includes SEQ ID NO:10's Amino acid sequence.

21. the Humanized anti-human CXCR3 antibody of claim 17, wherein the area the human IgG1 Fc includes SEQ ID NO:11's Amino acid sequence.

22. the method for claim 16, wherein the VH includes the amino acid sequence of SEQ ID NO:18, and the VL includes The amino acid sequence of SEQ ID NO:22.

23. the Humanized anti-human CXCR3 antibody of claim 22, wherein the area the human IgG1 Fc includes the ammonia of SEQ ID NO:2 Base acid sequence.

24. the Humanized anti-human CXCR3 antibody of claim 22, wherein the area the human IgG1 Fc includes the ammonia of SEQ ID NO:9 Base acid sequence.

25. the Humanized anti-human CXCR3 antibody of claim 22, wherein the area the human IgG1 Fc includes SEQ ID NO:10's Amino acid sequence.

26. the Humanized anti-human CXCR3 antibody of claim 22, wherein the area the human IgG1 Fc includes SEQ ID NO:11's Amino acid sequence.

27. the method for claim 16, wherein

A) HC includes the amino acid sequence of SEQ ID NO:25, and the LC includes the amino acid of SEQ ID NO:135 Sequence,

B) HC includes the amino acid sequence of SEQ ID NO:26, and the LC includes the amino acid of SEQ ID NO:135 Sequence,

C) HC includes the amino acid sequence of SEQ ID NO:27, and the LC includes the amino acid of SEQ ID NO:135 Sequence,

D) HC includes the amino acid sequence of SEQ ID NO:139, and the LC includes the amino acid of SEQ ID NO:135 Sequence,

E) HC includes the amino acid sequence of SEQ ID NO:31, and the LC includes the amino acid of SEQ ID NO:137 Sequence,

F) HC includes the amino acid sequence of SEQ ID NO:32, and the LC includes the amino acid of SEQ ID NO:137 Sequence,

G) HC includes the amino acid sequence of SEQ ID NO:32, and the LC includes the amino acid of SEQ ID NO:137 Sequence,

H) HC includes the amino acid sequence of SEQ ID NO:140, and the HC includes the amino acid of SEQ ID NO:137 Sequence.

28. the method for any one of claim 16-27, wherein the HC includes the human IgG1 with reduced fucose content The area Fc.

29. the method for any one of claim 16-27, wherein the antibody is generated in following host cell, the place Chief cell is cultivated in the presence of glycosylation inhibitor.

30. the method for claim 29, wherein the glycosylation inhibitor is base husband's alkali.

31. a kind of method for treating leucoderma comprising providing will be comprising heavy chain (HC) and tool with heavy chain variable region (VH) There is the Humanized anti-human CXCR3 antibody of the light chain (LC) of light chain variable region (VL) to give to the explanation of subject in need thereof Book, wherein

A) VH and VL include be respectively selected from SEQ ID NO:24/20,22/18,80/130,81/24,82/130,82/24, 83/126、83/130、83/24、84/126、84/24、85/24、86/126、87/126、87/133、87/24、88/24、89/ 24、90/126、91/126、91/130、92/130、92/24、93/126、94/126、95/126、96/126、97/126、98/ 126、99/126、100/126、101/126、102/126、103/126、104/126、105/126、106/126、107/126、 108/126、109/126、110/126、111/126、121/24、113/130、113/24、114/130、115/126、115/ 130、115/24、116/126、116/130、116/24、117/126、118/126、20/123、20/124、20/125、20/ 126、20/127、20/128、20/129、20/130、20/131、20/132、20/133、20/134、119/126、122/126、 With the amino acid sequence of 120/130 sequence pair, and wherein

B) heavy chain include the area human IgG1 Fc, the area the human IgG1 Fc have selected from SEQ ID NO:2,3,4,5,6,7,8, 9,10 and 11 amino acid sequence.

32. it is a kind of for treating the kit of leucoderma, it includes

A) Humanized anti-human CXCR3 antibody, the Humanized anti-human CXCR3 antibody include the heavy chain with heavy chain variable region (VH) (HC) and with light chain variable region (VL) light chain (LC), wherein

I) VH and VL include be respectively selected from SEQID NO:24/20,22/18,80/130,81/24,82/130,82/24, 83/126、83/130、83/24、84/126、84/24、85/24、86/126、87/126、87/133、87/24、88/24、89/ 24、90/126、91/126、91/130、92/130、92/24、93/126、94/126、95/126、96/126、97/126、98/ 126、99/126、100/126、101/126、102/126、103/126、104/126、105/126、106/126、107/126、 108/126、109/126、110/126、111/126、121/24、113/130、113/24、114/130、115/126、115/ 130、115/24、116/126、116/130、116/24、117/126、118/126、20/123、20/124、20/125、20/ 126、20/127、20/128、20/129、20/130、20/131、20/132、20/133、20/134、119/126、122/126、 With the amino acid sequence of 120/130 sequence pair, and wherein

Ii) HC also includes the area human IgG1 Fc, the area the human IgG1 Fc include selected from SEQ ID NO:2,3,4,5,6,7, 8,9,10 or 11 amino acid sequence；And

B) the Humanized anti-human CXCR3 is given to the specification of human experimenter in need thereof.