High Affinity Human Antibodies To Human Il-4 Receptor

Abstract

The present invention provides nucleic acid molecules that encode antibodies or antigen-binding fragments thereof, which specifically bind human interleukin-4 receptor (IL-4R). Also provided are expression vectors comprising nucleic acid molecule that encode anti-IL-4R antibodies, host cells comprising the expression vectors, and methods of producing anti-IL-4R antibodies or antigen-binding fragments thereof comprising growing the host cells under conditions permitting production of the antibody or fragment, and recovering the antibody or fragment so produced.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application is a continuation of Ser. No. 14/254,369, filed on Apr. 16, 2014, which is a continuation of Ser. No. 13/678,650, filed Nov. 16, 2012, issued as U.S. Pat. No. 8,735,095, which is a divisional of U.S. patent application Ser. No. 13/287,151, filed on Nov. 2, 2011, issued as U.S. Pat. No. 8,337,839, which is a divisional of U.S. patent application Ser. No. 12/556,605, filed on Sep. 10, 2009, issued as U.S. Pat. No. 8,075,887, which is a divisional of U.S. patent application Ser. No. 12/260,307, filed on Oct. 29, 2008, issued as U.S. Pat. No. 7,608,693, which is a continuation-in-part of U.S. patent application Ser. No. 11/906,517, filed on Oct. 2, 2007, issued as U.S. Pat. No. 7,605,237, which claims the benefit under 35 USC .sctn. 119(e) of U.S. Provisional Application No. 60/848,694, filed on Oct. 2, 2006, and U.S. Provisional Application No. 60/957,738, filed on Aug. 24, 2007, the contents of which are herein specifically incorporated by reference in their entireties.

BACKGROUND

[0002] Interleukin-4 (IL-4, also known as B cell stimulating factor or BSF-1) was originally characterized by its ability to stimulate the proliferation of B cells in response to low concentrations of antibodies directed to surface immunoglobulin. IL-4 has been shown to possess a broad spectrum of biological activities, including growth stimulation of T cells, mast cells, granulocytes, megakaryocytes and erythrocytes. IL-4 induces the expression of class II major histocompatibility complex molecules in resting B cells, and enhances the secretion of IgE and IgG1 isotypes by stimulated B cells.

[0003] The biological activities of IL-4 are mediated by specific cell surface receptors for IL-4. Human IL-4 receptor alpha (hIL-4R) (SEQ ID NO:274) is described in, for example, U.S. Pat. Nos. 5,599,905, 5,767,065, and 5,840,869. Antibodies to hIL-4R are described in U.S. Pat. Nos. 5,717,072 and 7,186,809.

[0004] Methods to produce antibodies useful as human therapeutics include generating chimeric antibodies and humanized antibodies (see, for example, U.S. Pat. No. 6,949,245). See, for example, WO 94/02602 and U.S. Pat. No. 6,596,541 (both of which publications are herein specifically incorporated by reference) describing methods of generating nonhuman transgenic mice capable of producing human antibodies.

[0005] Methods for using antibodies to hIL-4R are described in U.S. Pat. Nos. 5,714,146; 5,985,280; and 6,716,587.

BRIEF SUMMARY OF THE INVENTION

[0006] In a first aspect, the invention provides human antibodies, preferably recombinant human antibodies, that specifically bind human interleukin-4 receptor (hIL-4R). The human antibodies are characterized by binding to hIL-4R with high affinity and by the ability to neutralize hIL-4 activity. In specific embodiments, the human antibodies are capable of blocking hIL-13/hIL-13R1 complex binding to hIL-4R, and thus inhibit signaling by hIL-13. The antibodies can be full-length (for example, an IgG1 or IgG4 antibody) or may comprise only an antigen-binding portion (for example, a Fab, F(ab').sub.2 or scFv fragment), and may be modified to effect functionality, e.g., to eliminate residual effector functions (Reddy et al. (2000) J. Immunol. 164:1925-1933).

[0007] In a general embodiment, the invention provides an antibody or antigen-binding fragment thereof, that specifically binds hIL-4R (SEQ ID NO:274) with a K.sub.D of about 300 pM or less, as measured by surface plasmon resonance in a monomeric or dimeric assay. In a more specific embodiment, the antibody or antigen-binding portion thereof exhibits a K.sub.D of about 200 pM or less, about 150 or less, about 100 pM or less, or about 50 pM. In various embodiments, the antibody or antigen-binding fragment blocks hIL-4 activity with an IC.sub.50 of about 100 pM or less, as measured by luciferase bioassay. In more specific embodiments, the antibody or antigen-binding fragment exhibits an IC.sub.50 of about 50 pM or less, about 30 pM or less, or about 25 pM or less, as measured by STAT6 luciferase bioassay. In various embodiments, the antibody or antigen-binding fragment blocks hIL-13 activity with an IC.sub.50 of about 100 pM or less, about 90 pM or less, about 50 pM or less, or about 20 pM or less, as measured by STAT6 luciferase bioassay.

[0008] In a second aspect, the antibody of the invention comprises a heavy chain variable region (HCVR) sequence selected from the group consisting of SEQ ID NO:2, 18, 22, 26, 42, 46, 50, 66, 70, 74, 90, 94, 98, 114, 118, 122, 138, 142, 146, 162, 166, 170, 186, 190, 194, 210, 214, 218, 234, 238, 242, 258 and 262, or a substantially similar sequence thereof.

[0009] In a third aspect, the antibody of the invention comprises a light chain variable region (LCVR) sequence selected from the group consisting of SEQ ID NO:10, 20, 24, 34, 44, 48, 58, 68, 72, 82, 92, 96, 106, 116, 120, 130, 140, 144, 154, 164, 168, 178, 188, 192, 202, 212, 216, 226, 236, 240, 250, 260 and 264, or a substantially similar sequence thereof.

[0010] In one embodiment, the antibody or antibody fragment of the invention comprises HCVR and LCVR sequence pairs (HCVR/LCVR) selected from the group consisting of SEQ ID NO: 2/10, 18/20, 22/24, 26/34, 42/44, 46/48, 50/58, 66/68, 70/72, 74/82, 90/92, 94/96, 98/106, 114/116, 118/120, 122/130, 138/140, 142/144, 146/154, 162/164, 166/168, 170/178, 186/188, 190/192, 194/202, 210/212, 214/216, 218/226, 234/236, 238/240, 242/250, 258/260 and 262/264. In a preferred embodiment, the antibody or antibody fragment comprise HCVR/LCVR sequence pairs SEQ ID NO:162/164, 210/212 and 18/20. In an even more preferred embodiment, the antibody or antibody fragment comprises HCVR/LCVR SEQ ID NO:162/164.

[0011] In a fourth aspect, the invention provides nucleic acid molecules encoding an HCVR, wherein the nucleic acid molecule is a nucleotide sequence selected from the group consisting of SEQ ID NO: 1, 17, 21, 25, 41, 45, 49, 65, 69, 73, 89, 93, 97, 113, 117, 121, 137, 141, 145, 161, 165, 169, 185, 189, 193, 209, 213, 217, 233, 237, 241, 257 and 261, or a substantially identical sequence having at least 95% homology thereof.

[0012] In a fifth aspect, the invention provides nucleic acid molecules encoding a LCVR, wherein the nucleic acid molecule is a sequence selected from the group consisting of SEQ ID NO: 9, 19, 23, 33, 43, 47, 57, 67, 71, 81, 91, 95, 105, 115, 119, 129, 139, 143, 153, 163, 167, 177, 187, 191, 201, 211, 215, 225, 235, 239, 249, 259 and 263, or a substantially identical sequence having at least 95% homology thereof.

[0013] In one embodiment, the antibody of the invention comprises a HCVR and LCVR encoded by a nucleotide sequence pairs selected from the group consisting of SEQ ID NO: 1/9, 17/19, 21/22, 25/33, 41/43, 45/47, 49/57, 65/67, 69/71, 73/81, 89/91, 93/95, 97/105, 113/115, 117/119, 121/129, 137/139, 141/143, 145/153, 161/163, 165/167, 169/177, 185/187, 189/191, 193/201, 209/211, 213/215, 217/225, 233/235, 237/239, 241/249, 257/259 and 261/263. In a preferred embodiment, the antibody or antibody fragment comprise HCVR/LCVR sequences encoded by nucleic acid sequences selected from SEQ ID NO:161/163, 209/211 and 17/19. In an even more preferred embodiment, the antibody or antibody fragment comprises HCVR/LCVR encoded by nucleic acid sequences SEQ ID NO:161/163.

[0014] In a sixth aspect, the invention provides an antibody or antigen-binding fragment comprising a HCDR3 and a LCDR3, wherein the HCDR3 domain selected from the group consisting of SEQ ID NO:8, 32, 56, 80, 104, 128, 152, 176, 200, 224 and 248; and the LCDR3 domain selected from the group consisting of SEQ ID NO:16, 40, 64, 88, 112, 136, 160, 184, 208, 232 and 256. In a preferred embodiment, the HCDR3/LCDR3 sequences are SEQ ID NO:152/160, 8/16 or 200/208. In an even more preferred embodiment, the HCDR3 and LCDR3 sequences are SEQ ID NO:152 and 160.

[0015] In a further embodiment, the antibody or antibody fragment further comprises a HCDR1 sequence selected from the group consisting of SEQ ID NO:4, 28, 52, 76, 100, 124, 148, 172, 196, 220 and 244, or a substantially similar sequence thereof; a HCDR2 sequence selected from the group consisting of SEQ ID NO:6, 30, 54, 78, 102, 126, 150, 174, 198, 222 and 246, or a substantially similar sequence thereof; a HCDR3 sequence selected from the group consisting of SEQ ID NO:8, 32, 56, 80, 104, 128, 152, 176, 200, 224 and 248, or a substantially similar sequence thereof; a LCDR1 sequence selected from the group consisting of SEQ ID NO:12, 36, 60, 84, 108, 132, 156, 180, 204, 228 and 252, or a substantially similar sequence thereof; a LCDR2 sequence selected from the group consisting of SEQ ID NO:14, 38, 62, 86, 110, 134, 158, 182, 206, 230 and 252, or a substantially similar sequence thereof; and a LCDR3 sequence selected from the group consisting of SEQ ID NO:16, 40, 64, 88, 112, 136, 160, 184, 208, 232 and 256 or a substantially similar sequences thereof. In a preferred embodiment, the antibody or antigen-binding fragment comprise HCDR sequences SEQ ID NO:148, 150 and 152 and LCDR sequences SEQ ID NO:156, 158 and 160; HCDR sequences SEQ ID NO:4, 6 and 8 and LCDR sequences SEQ ID NO:12, 14 and 16; and HCDR sequences SEQ ID NO:196, 198 and 200 and LCDR sequences SEQ ID NO:204, 206 and 208.

[0016] In a seventh aspect, the invention features a human antibody or antibody fragment comprising a HCDR3 and LCDR3, wherein the HCDR3 is encoded by a nucleotide sequence selected from the group consisting of SEQ ID NO:7, 31, 55, 79, 103, 127, 151, 175, 199, 223 and 247, or a substantially identical sequence having at least 95% homology thereof; and the LCDR3 is encoded by a nucleotide sequence selected from the group consisting of SEQ ID NO:15, 39, 63, 87, 111, 135, 159, 183, 207, 231 and 255, or a substantially identical sequence having at least 95% homology thereof.

[0017] In a further embodiment, the invention features a human antibody or antibody fragment comprising a HCDR1 domain encoded by a nucleotide sequence selected from the group consisting of SEQ ID NO:3, 27, 51, 75, 99, 123, 147, 171, 195, 219 and 243, or a substantially identical sequence having at least 95% homology thereof; a HCDR2 domain encoded by a nucleotide sequence selected from the group consisting of SEQ ID NO:5, 29, 53, 77, 101, 125, 149, 173, 197, 221 and 245, or a substantially identical sequence having at least 95% homology thereof; a HCDR3 domain encoded by a nucleotide sequence selected from the group consisting of SEQ ID NO: 7, 31, 55, 79, 103, 127, 151, 175, 199, 223 and 247, or a substantially similar sequence having at least 95% homology thereof; a LCDR1 domain encoded by a nucleotide sequence selected from the group consisting of SEQ ID NO:11, 35, 59, 83, 107, 131, 155, 179, 203, 227 and 251, or a substantially similar sequence having at least 95% homology thereof; a LCDR2 domain encoded by a nucleotide sequence selected from the group consisting of SEQ ID NO:13, 37, 61, 85, 109, 133, 157, 181, 205, 229 and 253, or a substantially similar sequence having at least 95% homology thereof; and LCDR3 domain encoded by a nucleotide sequence selected from the group consisting of SEQ ID NO: 15, 39, 63, 87, 111, 135, 159, 183, 207, 231 and 255, or a substantially similar sequence having at least 95% homology thereof. In a preferred embodiment, the antibody or antigen-binding fragment comprise HCDR and LCDR sequences encoded by nucleotide sequences SEQ ID NO:147, 149, 151, 155, 157 and 159; 195, 197, 199, 203, 205 and 207; and 3, 5, 7, 11, 13 and 15.

[0018] In a specific embodiment, the anti-hIL-4R antibody or antigen-binding fragment of the invention comprises HCVR comprising the amino acid sequence shown in SEQ ID NO:162 and LCVR comprising the amino acid sequence shown in SEQ ID NO:164, and is characterized by a K.sub.D of about 100 pM or less (monomeric substrate) or 70 pM or less (dimeric substrate); a K.sub.D of about 160 pM or less (monomeric substrate) or 40 pM or less (dimeric substrate) at 25.degree. C. and 37.degree. C., respectively; and an IC.sub.50 of about 10 pM or less (25 pM dimer substrate) or about 100 pM or less (200 pM monomer substrate), which is capable of blocking both hIL-4 and hIL-13 activity with an IC.sub.50 of about 30 pM or less (as measured by bioassay) and cross-reacts with monkey IL-4R.

[0019] In a specific embodiment, the anti-hIL-4R antibody or antigen-binding fragment of the invention comprises HCVR comprising the amino acid sequence shown in SEQ ID NO:18 and LCVR comprising the amino acid sequence shown in SEQ ID NO:20, and is characterized by a K.sub.D of about 450 pM or less (monomeric or dimeric substrate); and an IC.sub.50 of about 40 pM or less (25 pM dimer substrate) or about 100 pM or less (200 pM monomer substrate), which is capable of blocking both hIL-4 and hIL-13 activity with an IC.sub.50 of about 100 pM or less (as measured by bioassay).

[0020] In a specific embodiment, the anti-hIL-4R antibody or antigen-binding fragment of the invention comprises HCVR comprising the amino acid sequence shown in SEQ ID NO:210 and LCVR comprising the amino acid sequence shown in SEQ ID NO:212, and is characterized by a K.sub.D of about 50 pM or less (monomeric substrate) or 30 pM or less (dimeric substrate); a K.sub.D of about 200 pM or less (monomeric substrate) or 40 pM or less (dimeric substrate) at 25.degree. C. and 37.degree. C., respectively; and an IC.sub.50 of about 10 pM or less (25 pM dimer substrate) or about 90 pM or less (200 pM monomer substrate), which is capable of blocking both hIL-4 and hIL-13 activity with an IC.sub.50 of about 25 pM or less (as measured by bioassay) and does not cross-reacts with monkey IL-4R.

[0021] In a eighth aspect, the invention features an antibody or antigen-binding fragment of an antibody that specifically binds hIL-4R, comprising three heavy chain and three light chain complementarity determining regions (HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, LCDR3), wherein the HCDR1 comprises an amino acid sequence of the formula X.sup.1-X.sup.2-X.sup.3-X.sup.4-X.sup.5-X.sup.6-X.sup.7-X.sup.8 (SEQ ID NO:265), wherein X.sup.1=Gly; X.sup.2=Phe; X.sup.3=Thr; X.sup.4=Phe; X.sup.5=Asp or Arg; X.sup.6=Asp or Ser; X.sup.7=Tyr; and X.sup.8=Ala or Gly; the HCDR2 comprises an amino acid sequence of the formula X.sup.1-X.sup.2-X.sup.3-X.sup.4-X.sup.5-X.sup.6-X.sup.7-X.sup.8 (SEQ ID NO:266), wherein X.sup.1=Ile or Leu, X.sup.2=Ser, X.sup.3=Gly, Tyr or Arg, X.sup.4=Ser, Asp or Thr, X.sup.5=Gly or Ser, X.sup.6=Gly, Ser or Val, X.sup.7=Ser or Asn, and X.sup.8=Thr, Lys or Ile; the HCDR3 comprises an amino acid sequence of the formula X.sup.1-X.sup.2-X.sup.3-X.sup.4-X.sup.5-X.sup.6-X.sup.7-X.sup.8-X.sup.9-X- .sup.10-X.sup.11-X.sup.12-X.sup.13-X.sup.14-X.sup.15-X.sup.16-X.sup.17-X.s- up.18 (SEQ ID NO:267) wherein X.sup.1=Ala, X.sup.2=Lys, X.sup.3=Asp, Glu or Trp, X.sup.4=Gly or Arg, X.sup.5=Leu, Thr or Arg, X.sup.6=Gly, Arg or Ser, X.sup.7=Ile or Gly, X.sup.8=Thr, Phe or Tyr, X.sup.9=Ile, Asp or Phe, X.sup.10=Arg, Tyr or Asp, X.sup.11=Pro, Tyr or absent, X.sup.12=Arg or absent, X.sup.13=Tyr or absent, X.sup.14=Tyr or absent, X.sup.15=Gly or absent, X.sup.16=Leu or absent, X.sup.17=Asp or absent, and X.sup.18=Val or absent; the LCDR1 comprises an amino acid sequence of the formula X.sup.1-X.sup.2-X.sup.3-X.sup.4-X.sup.5-X.sup.6-X.sup.7-X.sup.8-X- .sup.9-X.sup.10-X.sup.11 (SEQ ID NO:268) wherein X.sup.1=Gln, X.sup.2=Asp, Ser or Val, X.sup.3=Ile or Leu, X.sup.4=Ser, Leu or Asn, X.sup.5=Asn, Tyr or Ile, X.sup.6=Trp, Ser or Tyr; X.sup.7=Ile or absent; X.sup.8=Gly or absent; X.sup.9=Tyr or absent; X.sup.10=Asn or absent; and X.sup.11=Tyr or absent; the LCDR2 comprises an amino acid sequence of the formula X.sup.1-X.sup.2-X.sup.3 (SEQ ID NO:269) wherein X.sup.1=Leu, Ala or Val, X.sup.2=Ala or Gly, and X.sup.3=Ser; and the LCDR3 comprises an amino acid sequence of the formula X.sup.1-X.sup.2-X.sup.3-X.sup.4-X.sup.5-X.sup.6-X.sup.7-X.sup.8-X.sup.9 (SEQ ID NO:270) wherein X.sup.1=Gln or Met, X.sup.2=Gln, X.sup.3=Ala or Tyr, X.sup.4=Leu or Asn, X.sup.5=Gln or Ser, X.sup.6=Thr, Phe or His, X.sup.7=Pro, X.sup.8=Tyr, Ile or Trp, and X.sup.9=Thr.

[0022] In a more specific embodiment, the HCDR1 comprises an amino acid sequence of the formula X.sup.1-X.sup.2-X.sup.3-X.sup.4-X.sup.5-X.sup.6-X.sup.7-X.sup.8 (SEQ ID NO:265), wherein X.sup.1=Gly; X.sup.2=Phe; X.sup.3=Thr; X.sup.4=Phe; X.sup.5=Arg; X.sup.6=Asp or Ser; X.sup.7=Tyr; and X.sup.8=Ala or Gly; the HCDR2 comprises an amino acid sequence of the formula X.sup.1-X.sup.2-X.sup.3-X.sup.4-X.sup.5-X.sup.6-X.sup.7-X.sup.8 (SEQ ID NO:266), wherein X.sup.1=Ile, X.sup.2=Ser, X.sup.3=Gly or Tyr, X.sup.4=Ser or Thr, X.sup.5=Gly, X.sup.6=Gly or Ser, X.sup.7=Asn, and X.sup.8=Thr or Lys; the HCDR3 comprises an amino acid sequence of the formula X.sup.1-X.sup.2-X.sup.3-X.sup.4-X.sup.5-X.sup.6-X.sup.7-X.sup.8-X- .sup.9-X.sup.10-X.sup.11-X.sup.12-X.sup.13-X.sup.14-X.sup.15-X.sup.16-X.su- p.17-X.sup.18 (SEQ ID NO:267) wherein X.sup.1=Ala, X.sup.2=Lys, X.sup.3=Asp or Glu, X.sup.4=Gly or Arg, X.sup.5=Leu or Arg, X.sup.6=Gly or Ser, X.sup.7=Ile or Gly, X.sup.8=Thr or Phe, X.sup.9=Ile or Asp, X.sup.10=Arg or Tyr, X.sup.11=Pro or absent, X.sup.12=Arg or absent, X.sup.13=Tyr or absent, X.sup.14=Tyr or absent, X.sup.15=Gly or absent, X.sup.16=Leu or absent, X.sup.17=Asp or absent, and X.sup.18=Val or absent; the LCDR1 comprises an amino acid sequence of the formula X.sup.1-X.sup.2-X.sup.3-X.sup.4-X.sup.5-X.sup.6-X.sup.7-X.sup.8-X.sup.9-X- .sup.10-X.sup.11 (SEQ ID NO:268) wherein X.sup.1=Gln, X.sup.2=Ser or Val, X.sup.3=Ile or Leu, X.sup.4=Leu or Asn, X.sup.5=Asn or Tyr, X.sup.6=Ser or Tyr; X.sup.7=Ile or absent; X.sup.8=Gly or absent; X.sup.9=Tyr or absent; X.sup.10=Asn or absent; and X.sup.11=Tyr or absent; the LCDR2 comprises an amino acid sequence of the formula X.sup.1-X.sup.2-X.sup.3 (SEQ ID NO:269) wherein X.sup.1=Leu or Ala, X.sup.2=Ala or Gly, and X.sup.3=Ser; and the LCDR3 comprises an amino acid sequence of the formula X.sup.1-X.sup.2-X.sup.3-X.sup.4-X.sup.5-X.sup.6-X.sup.7-X.sup.8-X- .sup.9 (SEQ ID NO:270) wherein X.sup.1=Gln or Met, X.sup.2=Gln, X.sup.3=Ala or Tyr, X.sup.4=Leu or Asn, X.sup.5=Gln or Ser, X.sup.6=Thr or His, X.sup.7=Pro, X.sup.8=Tyr or Trp, and X.sup.9=Thr.

[0023] In another more specific embodiment, the HCDR1 comprises an amino acid sequence of the formula X.sup.1-X.sup.2-X.sup.3-X.sup.4-X.sup.5-X.sup.6-X.sup.7-X.sup.8 (SEQ ID NO:265), wherein X.sup.1=Gly; X.sup.2=Phe; X.sup.3=Thr; X.sup.4=Phe; X.sup.5=Asp or Arg; X.sup.6=Asp; X.sup.7=Tyr; and X.sup.8=Ala; the HCDR2 comprises an amino acid sequence of the formula X.sup.1-X.sup.2-X.sup.3-X.sup.4-X.sup.5-X.sup.6-X.sup.7-X.sup.8 (SEQ ID NO:266), wherein X.sup.1=Ile or Leu, X.sup.2=Ser, X.sup.3=Gly or Arg, X.sup.4=Ser or Thr, X.sup.5=Gly or Ser, X.sup.6=Gly or Val, X.sup.7=Ser or Asn, and X.sup.8=Thr or Ile; the HCDR3 comprises an amino acid sequence of the formula X.sup.1-X.sup.2-X.sup.3-X.sup.4-X.sup.5-X.sup.6-X.sup.7-X.sup.8-X.sup.9-X- .sup.10-X.sup.11-X.sup.12-X.sup.13-X.sup.14-X.sup.15-X.sup.16-X.sup.17-X.s- up.18 (SEQ ID NO:267) wherein X.sup.1=Ala, X.sup.2=Lys, X.sup.3=Asp or Trp, X.sup.4=Gly or Arg, X.sup.5=Leu or Thr, X.sup.6=Arg or Ser, X.sup.7=Ile or Gly, X.sup.8=Thr or Tyr, X.sup.9=Ile or Phe, X.sup.10=Arg or Asp, X.sup.11=Pro, Tyr or absent, X.sup.12=Arg or absent, X.sup.13=Tyr or absent, X.sup.14=Tyr or absent, X.sup.15=Gly or absent, X.sup.16=Leu or absent, X.sup.17=Asp or absent, and X.sup.18=Val or absent; the LCDR1 comprises an amino acid sequence of the formula X.sup.1-X.sup.2-X.sup.3-X.sup.4-X.sup.5-X.sup.6-X.sup.7-X.sup.8-X.sup.9-X- .sup.10-X.sup.11 (SEQ ID NO:268) wherein X.sup.1=Gln, X.sup.2=Asp or Ser, X.sup.3=Ile or Leu, X.sup.4=Ser or Leu, X.sup.5=Tyr or Ile, X.sup.6=Trp or Ser; X.sup.7=Ile or absent; X.sup.8=Gly or absent; X.sup.9=Tyr or absent; X.sup.10=Asn or absent; and X''=Tyr or absent; the LCDR2 comprises an amino acid sequence of the formula X.sup.1-X.sup.2-X.sup.3 (SEQ ID NO:269) wherein X.sup.1=Leu or Val, X.sup.2=Ala or Gly, and X.sup.3=Ser; and the LCDR3 comprises an amino acid sequence of the formula X.sup.1-X.sup.2-X.sup.3-X.sup.4-X.sup.5-X.sup.6-X.sup.7-X.sup.8-X- .sup.9 (SEQ ID NO:270) wherein X.sup.1=Gln or Met, X.sup.2=Gln, X.sup.3=Ala, X.sup.4=Leu or Asn, X.sup.5=Gln or Ser, X.sup.6=Thr or Phe, X.sup.7=Pro, X.sup.8=Tyr or Ile, and X.sup.9=Thr.

[0024] In a ninth aspect, the invention provides an antibody or antigen-binding fragment comprising HCDR1/HCDR2/HCDR3/LCDR1/LCDR2/LCDR3 sequences from a HCVR and LCVR pair, wherein the HCVR/LCVR sequences are selected from the group consisting of SEQ ID NO:162/164, 210/212 and 18/20. In a more specific embodiment, heavy and light chain CDR sequences are those contained in HCVR SEQ ID NO:162 and LCVR SEQ ID NO:164. In another more specific embodiment, heavy and light chain CDR sequences are those contained in HCVR SEQ ID NO:18 and LCVR SEQ ID NO:20. In yet another specific embodiment, heavy and light chain CDR sequences are those contained in HCVR SEQ ID NO:210 and LCVR SEQ ID NO:212.

[0025] The invention encompasses anti-hIL-4R antibodies having a modified glycosylation pattern. In some applications, modification to remove undesirable glycosylation sites may be useful, or an antibody lacking a fucose moiety present on the oligosaccharide chain, for example, to increase antibody dependent cellular cytotoxicity (ADCC) function (see Shield et al. (2002) JBC 277:26733). In other applications, modification of a galactosylation can be made in order to modify complement dependent cytotoxicity (CDC).

[0026] In a tenth aspect, the invention provides recombinant expression vectors carrying the nucleic acid molecules of the invention, and host cells into which such vectors have been included, as are methods of making the antibodies or antigen-binding fragments of the invention obtained by culturing the host cells of the invention. The host cell may be a prokaryotic or eukaryotic cell, preferably the host cell is an E. coli cell or a mammalian cell, such as a CHO cell.

[0027] In an eleventh aspect, the invention features a composition comprising a recombinant human antibody that specifically binds hIL-4R and an acceptable carrier.

[0028] In a twelfth aspect, the invention features methods for inhibiting hIL-4 activity using an antibody, or antigen-binding portion thereof, of the invention. In specific embodiments, the antibodies of the invention also block hIL-13/hIL-13R1 complex binding to hIL-4R. In one embodiment, the method comprises contacting hIL-4R with the antibody of the invention, or antigen-binding portion thereof, such that hIL-4 or hIL-4/hIL-13 activity is inhibited. In another embodiment, the method comprises administering an antibody of the invention, or antigen-binding portion thereof, to a human subject suffering from a disorder that is ameliorated by inhibition of hIL-4 or hIL-4/hIL-13 activity. The disorder treated is any disease or condition that is improved, ameliorated, inhibited or prevented by removal, inhibition or reduction of hIL-4 or hIL-4/hIL-13 activity.

[0029] IL-4 related disorders which are treated by the antibodies or antibody fragments of the invention include, for example, arthritis (including septic arthritis), herpetiformis, chronic idiopathic urticaria, scleroderma, hypertrophic scarring, Whipple's Disease, benign prostate hyperplasia, lung disorders, such as mild, moderate or severe asthma, inflammatory disorders such as inflammatory bowel disease, allergic reactions, Kawasaki disease, sickle cell disease, Churg-Strauss syndrome, Grave's disease, pre-eclampsia, Sjogren's syndrome, autoimmune lymphoproliferative syndrome, autoimmune hemolytic anemia, Barrett's esophagus, autoimmune uveitis, tuberculosis, and nephrosis.

[0030] Other objects and advantages will become apparent from a review of the ensuing detailed description.

DETAILED DESCRIPTION

[0031] Before the present methods are described, it is to be understood that this invention is not limited to particular methods, and experimental conditions described, as such methods and conditions may vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting, since the scope of the present invention will be limited only by the appended claims.

[0032] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, the preferred methods and materials are now described. All publications mentioned herein are incorporated herein by reference in their entirety.

Definitions

[0033] The term "human IL4R" (hIL-4R), as used herein, is intended to refer to a human cytokine receptor that specifically binds interleukin-4 (IL-4), IL-4R.alpha. (SEQ ID NO:274). The term "human interleukin-13" (hIL-13) refers to a cytokine that specifically binds IL-13 receptor, and "hIL-13/hIL-13R1 complex" refers to the complex formed by hIL-13 binding to hIL-13R1 complex, which complex binds hIL-4 receptor to initiate biological activity.

[0034] The term "antibody", as used herein, is intended to refer to immunoglobulin molecules comprising four polypeptide chains, two heavy (H) chains and two light (L) chains inter-connected by disulfide bonds. Each heavy chain comprises a heavy chain variable region (HCVR or VH) and a heavy chain constant region. The heavy chain constant region comprises three domains, CH1, CH2 and CH3. Each light chain comprises a light chain variable region (LCVR or VL) and a light chain constant region. The light chain constant region comprises one domain (CL1). The VH and VL regions can be further subdivided into regions of hypervariability, termed complementarity determining regions (CDR), interspersed with regions that are more conserved, termed framework regions (FR). Each VH and VL is composed of three CDRs and four FRs, arranged from amino-terminus to carboxy-terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4.

[0035] The term "antigen-binding portion" of an antibody (or simply "antibody portion" or "antibody fragment"), as used herein, refers to one or more fragments of an antibody that retain the ability to specifically bind to an antigen (e.g., hIL-4R). It has been shown that the antigen-binding function of an antibody can be performed by fragments of a full-length antibody. Examples of binding fragments encompassed within the term "antigen-binding portion" of an antibody include (i) a Fab fragment, a monovalent fragment consisting of the VL, VH, CL1 and CH1 domains; (ii) a F(ab').sub.2 fragment, a bivalent fragment comprising two F(ab)' fragments linked by a disulfide bridge at the hinge region; (iii) a Fd fragment consisting of the VH and CH1 domains; (iv) a Fv fragment consisting of the VL and VH domains of a single arm of an antibody, (v) a dAb fragment (Ward et al. (1989) Nature 241:544-546), which consists of a VH domain; and (vi) an CDR. Furthermore, although the two domains of the Fv fragment, VL and VH, are coded for by separate genes, they can be joined, using recombinant methods, by a synthetic linker that enables them to be made as a single contiguous chain in which the VL and VH regions pair to form monovalent molecules (known as single chain Fv (scFv); see e.g., Bird et al. (1988) Science 242:423-426; and Huston et al. (1988) Proc. Natl. Acad. Sci. USA 85:5879-5883. Such single chain antibodies are also intended to be encompassed within the term "antigen-binding portion" of an antibody. Other forms of single chain antibodies, such as diabodies, are also encompassed (see e.g., Holliger et al. (1993) Proc. Natl. Acad Sci. USA 90:6444-6448).

[0036] A "neutralizing" or "blocking" antibody, as used herein, is intended to refer to an antibody whose binding to hIL-4R results in inhibition of the biological activity of hIL-4 and/or hIL-13. This inhibition of the biological activity of hIL-4 and/or IL-13 can be assessed by measuring one or more indicators of hIL-4 and/or hIL-13 biological activity known to the art, such as hIL-4- and/or IL-13-induced cellular activation and hIL-4 binding to hIL-4R (see examples below).

[0037] A "CDR" or complementarity determining region is a region of hypervariability interspersed within regions that are more conserved, termed "framework regions" (FR). In different embodiments of the anti-hIL-4R antibody or fragment of the invention, the FRs may be identical to the human germline sequences, or may be naturally or artificially modified.

[0038] The term "surface plasmon resonance", as used herein, refers to an optical phenomenon that allows for the analysis of real-time interactions by detection of alterations in protein concentrations within a biosensor matrix, for example using the BIACORE.TM. system (Pharmacia Biosensor AB).

[0039] The term "epitope" is an antigenic determinant that interacts with a specific antigen binding site in the variable region of an antibody molecule known as a paratope. A single antigen may have more than one epitope. Epitopes may be either conformational or linear. A conformational epitope is produced by spatially juxtaposed amino acids from different segments of the linear polypeptide chain. A linear epitope is one produced by adjacent amino acid residues in a polypeptide chain. In certain circumstance, an epitope may include moieties of saccharides, phosphoryl groups, or sufonyl groups on the antigen.

[0040] The term "substantial identity" or "substantially identical," when referring to a nucleic acid or fragment thereof, indicates that, when optimally aligned with appropriate nucleotide insertions or deletions with another nucleic acid (or its complementary strand), there is nucleotide sequence identity in at least about 95%, and more preferably at least about 96%, 97%, 98% or 99% of the nucleotide bases, as measured by any well-known algorithm of sequence identity, such as FASTA, BLAST or Gap, as discussed below.

[0041] As applied to polypeptides, the term "substantial similarity" or "substantially similar" means that two peptide sequences, when optimally aligned, such as by the programs GAP or BESTFIT using default gap weights, share at least 95% sequence identity, even more preferably at least 98% or 99% sequence identity. Preferably, residue positions which are not identical differ by conservative amino acid substitutions. A "conservative amino acid substitution" is one in which an amino acid residue is substituted by another amino acid residue having a side chain (R group) with similar chemical properties (e.g., charge or hydrophobicity). In general, a conservative amino acid substitution will not substantially change the functional properties of a protein. In cases where two or more amino acid sequences differ from each other by conservative substitutions, the percent sequence identity or degree of similarity may be adjusted upwards to correct for the conservative nature of the substitution. Means for making this adjustment are well-known to those of skill in the art. See, e.g., Pearson (1994) Methods Mol. Biol. 24: 307-331, herein incorporated by reference. Examples of groups of amino acids that have side chains with similar chemical properties include (1) aliphatic side chains: glycine, alanine, valine, leucine and isoleucine; (2) aliphatic-hydroxyl side chains: serine and threonine; (3) amide-containing side chains: asparagine and glutamine; (4) aromatic side chains: phenylalanine, tyrosine, and tryptophan; (5) basic side chains: lysine, arginine, and histidine; (6) acidic side chains: aspartate and glutamate, and (7) sulfur-containing side chains are cysteine and methionine. Preferred conservative amino acids substitution groups are: valine-leucine-isoleucine, phenylalanine-tyrosine, lysine-arginine, alanine-valine, glutamate-aspartate, and asparagine-glutamine. Alternatively, a conservative replacement is any change having a positive value in the PAM250 log-likelihood matrix disclosed in Gonnet et al. (1992) Science 256: 1443 45, herein incorporated by reference. A "moderately conservative" replacement is any change having a nonnegative value in the PAM250 log-likelihood matrix.

[0042] Sequence similarity for polypeptides, which is also referred to as sequence identity, is typically measured using sequence analysis software. Protein analysis software matches similar sequences using measures of similarity assigned to various substitutions, deletions and other modifications, including conservative amino acid substitutions. For instance, GCG software contains programs such as Gap and Bestfit which can be used with default parameters to determine sequence homology or sequence identity between closely related polypeptides, such as homologous polypeptides from different species of organisms or between a wild type protein and a mutein thereof. See, e.g., GCG Version 6.1. Polypeptide sequences also can be compared using FASTA using default or recommended parameters, a program in GCG Version 6.1. FASTA (e.g., FASTA2 and FASTA3) provides alignments and percent sequence identity of the regions of the best overlap between the query and search sequences (Pearson (2000) supra). Another preferred algorithm when comparing a sequence of the invention to a database containing a large number of sequences from different organisms is the computer program BLAST, especially BLASTP or TBLASTN, using default parameters. See, e.g., Altschul et al. (1990) J. Mol. Biol. 215:403-410 and Altschul et al. (1997) Nucleic Acids Res. 25:3389-402, each of which is herein incorporated by reference.

Preparation of Human Antibodies

[0043] Methods for generating human antibodies include those described in, for example, U.S. Pat. No. 6,596,541, Green et al. (1994) Nature Genetics 7:13-21), U.S. Pat. No. 5,545,807, U.S. Pat. No. 6,787,637.

[0044] Rodents can be immunized by any method known in the art (see, for example, Harlow and Lane (1988) Antibodies: A Laboratory Manual 1988 Cold Spring Harbor Laboratory; Malik and Lillehoj (1994) Antibody Techniques, Academic Press, CA). Antibodies of the invention are preferably prepared with the use of VELOCIMMUNE.TM. technology (U.S. Pat. No. 6,596,541). A transgenic mouse in which the endogenous immunoglobulin heavy and light chain variable regions are replaced with the corresponding human variable regions is challenged with the antigen of interest, and lymphatic cells (such as B-cells) are recovered from the mice that express antibodies. The lymphatic cells may be fused with a myeloma cell line to prepare immortal hybridoma cell lines, and such hybridoma cell lines are screened and selected to identify hybridoma cell lines that produce antibodies specific to the antigen of interest. DNA encoding the variable regions of the heavy chain and light chain may be isolated and linked to desirable isotypic constant regions of the heavy chain and light chain. Such an antibody protein may be produced in a cell, such as a CHO cell. Alternatively, DNA encoding the antigen-specific chimeric antibodies or the variable regions of the light and heavy chains may be isolated directly from antigen-specific lymphocytes.

[0045] The DNA encoding the variable regions of the heavy and light chains of the antibody are isolated and operably linked to DNA encoding the human heavy and light chain constant regions. The DNA is then expressed in a cell capable of expressing the fully human antibody. In a specific embodiment, the cell is a CHO cell.

[0046] Antibodies may be therapeutically useful in blocking a ligand-receptor interaction or inhibiting receptor component interaction, rather than by killing cells through fixation of complement (complement-dependent cytotoxicity) (CDC) and participation antibody-dependent cell-mediated cytotoxicity (ADCC). The constant region of an antibody is important in the ability of an antibody to fix complement and mediate cell-dependent cytotoxicity. Thus, the isotype of an antibody may be selected on the basis of whether it is desirable for the antibody to mediate cytotoxicity.

[0047] Human immunoglobulins can exist in two forms that are associated with hinge heterogeneity. In one form, an immunoglobulin molecule comprises a stable four-chain construct of approximately 150-160 kDa in which the dimers are held together by an interchain heavy chain disulfide bond. In a second form, the dimers are not linked via interchain disulfide bonds and a molecule of about 75-80 kDa is formed composed of a covalently coupled light and heavy chain (half-antibody). These forms have been extremely difficult to separate, even after affinity purification. The frequency of appearance of the second form in various intact IgG isotypes is due to, but not limited to, structural differences associated with the hinge region isotype of the antibody. In fact, a single amino acid substitution in the hinge region of the human IgG4 hinge can significantly reduce the appearance of the second form (Angal et al. (1993) Molecular Immunology 30: 105) to levels typically observed using a human IgG1 hinge. The instant invention encompasses antibodies having one or more mutations in the hinge, CH2 or CH3 region that may be desirable, for example, in production, to improve the yield of the desired antibody form.

[0048] Initially, high affinity chimeric antibodies are isolated having a human variable region and a mouse constant region. As described below, the antibodies are characterized and selected for desirable characteristics, including binding affinity to hIL-4R, ability to block hIL-4 binding to hIL-4R, and/or selectivity for the human protein. The mouse constant regions are replaced with desired human constant regions to generate the fully human antibodies of the invention, for example wild-type or modified IgG4 or IgG1 (for example, SEQ ID NO:271, 272, 273). While the constant region selected may vary according to specific use, high affinity antigen-binding and target specificity characteristics reside in the variable region.

Epitope Mapping and Related Technologies

[0049] To screen for antibodies that bind to a particular epitope, a routine cross-blocking assay such as that described in Harlow and Lane supra can be performed. Other methods include alanine scanning mutants, peptide blots (Reineke (2004) Methods Mol Biol 248:443-63), or peptide cleavage analysis. In addition, methods such as epitope excision, epitope extraction and chemical modification of antigens can be employed (Tomer (2000) Protein Science: 9:487-496).

[0050] Modification-Assisted Profiling (MAP), also known as Antigen Structure-based Antibody Profiling (ASAP) is a method that categorizes large numbers of monoclonal antibodies (mAbs) directed against the same antigen according to the similarities of the binding profile of each antibody to chemically or enzymatically modified antigen surfaces (US Patent Application Publication No. 2004/0101920, herein specifically incorporated by reference in its entirety). Each category may reflect a unique epitope either distinctly different from, or partially overlapping with, an epitope represented by another category. This technology allows rapid filtering of genetically identical antibodies, such that characterization can be focused on genetically distinct antibodies. When applied to hybridoma screening, MAP may facilitate identification of rare hybridoma clones with desired characteristics. MAP may be used to sort the hIL-4R antibodies of the invention into groups of antibodies binding different epitopes.

[0051] Agents useful for altering the structure of the immobilized antigen are enzymes, such as, for example, proteolytic enzymes and chemical agents. The antigen protein may be immobilized on either biosensor chip surfaces or polystyrene beads. The latter can be processed with, for example, an assay such as a multiplex LUMINEX.TM. detection assay (Luminex Corp., TX). Because of the capacity of LUMINEX.TM. to handle multiplex analysis with up to 100 different types of beads, LUMINEX.TM. provides almost unlimited antigen surfaces with various modifications, resulting in improved resolution in antibody epitope profiling over a biosensor assay.

Bispecifics

[0052] The antibodies of the present invention may be monospecific, bispecific, or multispecific. Multispecific antibodies may be specific for different epitopes of one target polypeptide or may contain antigen-binding domains specific for more than one target polypeptide. See, e.g., Tutt et al. (1991) J. Immunol. 147:60-69. The human anti-IL-4R antibodies can be linked to or co-expressed with another functional molecule, e.g., another peptide or protein. For example, an antibody or fragment thereof can be functionally linked (e.g., by chemical coupling, genetic fusion, noncovalent association or otherwise) to one or more other molecular entities, such as another antibody or antibody fragment, to produce a bispecific or a multispecific antibody with a second binding specificity.

Therapeutic Administration and Formulations

[0053] The invention provides therapeutic compositions comprising the anti-IL-4R antibodies or antigen-binding fragments thereof of the present invention. The administration of therapeutic compositions in accordance with the invention will be administered with suitable carriers, excipients, and other agents that are incorporated into formulations to provide improved transfer, delivery, tolerance, and the like. A multitude of appropriate formulations can be found in the formulary known to all pharmaceutical chemists: Remington's Pharmaceutical Sciences, Mack Publishing Company, Easton, Pa. These formulations include, for example, powders, pastes, ointments, jellies, waxes, oils, lipids, lipid (cationic or anionic) containing vesicles (such as LIPOFECTIN.TM.), DNA conjugates, anhydrous absorption pastes, oil-in-water and water-in-oil emulsions, emulsions carbowax (polyethylene glycols of various molecular weights), semi-solid gels, and semi-solid mixtures containing carbowax. See also Powell et al. "Compendium of excipients for parenteral formulations" PDA (1998) J Pharm Sci Technol 52:238-311.

[0054] The dose may vary depending upon the age and the size of a subject to be administered, target disease, conditions, route of administration, and the like. When the antibody of the present invention is used for treating various conditions and diseases associated with IL-4R, in an adult patient, it is advantageous to intravenously administer the antibody of the present invention normally at a single dose of about 0.01 to about 20 mg/kg body weight, more preferably about 0.02 to about 7, about 0.03 to about 5, or about 0.05 to about 3 mg/kg body weight. Depending on the severity of the condition, the frequency and the duration of the treatment can be adjusted.

[0055] Various delivery systems are known and can be used to administer the pharmaceutical composition of the invention, e.g., encapsulation in liposomes, microparticles, microcapsules, recombinant cells capable of expressing the mutant viruses, receptor mediated endocytosis (see, e.g., Wu et al. (1987) J. Biol. Chem. 262:4429-4432). Methods of introduction include, but are not limited to, intradermal, intramuscular, intraperitoneal, intravenous, subcutaneous, intranasal, epidural, and oral routes. The composition may be administered by any convenient route, for example by infusion or bolus injection, by absorption through epithelial or mucocutaneous linings (e.g., oral mucosa, rectal and intestinal mucosa, etc.) and may be administered together with other biologically active agents. Administration can be systemic or local.

[0056] The pharmaceutical composition can be also delivered in a vesicle, in particular a liposome (see Langer (1990) Science 249:1527-1533; Treat et al. (1989) in Liposomes in the Therapy of Infectious Disease and Cancer, Lopez Berestein and Fidler (eds.), Liss, New York, pp. 353-365; Lopez-Berestein, ibid., pp. 317-327; see generally ibid.

[0057] In certain situations, the pharmaceutical composition can be delivered in a controlled release system. In one embodiment, a pump may be used (see Langer, supra; Sefton (1987) CRC Crit. Ref. Biomed. Eng. 14:201). In another embodiment, polymeric materials can be used (see Medical Applications of Controlled Release, Langer and Wise (eds.), CRC Pres., Boca Raton, Fla. (1974). In yet another embodiment, a controlled release system can be placed in proximity of the composition's target, thus requiring only a fraction of the systemic dose (see, e.g., Goodson, in Medical Applications of Controlled Release, supra, vol. 2, pp. 115-138, 1984). Other controlled release systems are discussed in the review by Langer (1990) Science 249:1527-1533.

[0058] The injectable preparations may include dosage forms for intravenous, subcutaneous, intracutaneous and intramuscular injections, drip infusions, etc. These injectable preparations may be prepared by methods publicly known. For example, the injectable preparations may be prepared, e.g., by dissolving, suspending or emulsifying the antibody or its salt described above in a sterile aqueous medium or an oily medium conventionally used for injections. As the aqueous medium for injections, there are, for example, physiological saline, an isotonic solution containing glucose and other auxiliary agents, etc., which may be used in combination with an appropriate solubilizing agent such as an alcohol (e.g., ethanol), a polyalcohol (e.g., propylene glycol, polyethylene glycol), a nonionic surfactant [e.g., polysorbate 80, HCO-50 (polyoxyethylene (50 mol) adduct of hydrogenated castor oil)], etc. As the oily medium, there are employed, e.g., sesame oil, soybean oil, etc., which may be used in combination with a solubilizing agent such as benzyl benzoate, benzyl alcohol, etc. The injection thus prepared is preferably filled in an appropriate ampoule.

[0059] Advantageously, the pharmaceutical compositions for oral or parenteral use described above are prepared into dosage forms in a unit dose suited to fit a dose of the active ingredients. Such dosage forms in a unit dose include, for example, tablets, pills, capsules, injections (ampoules), suppositories, etc. The amount of the aforesaid antibody contained is generally about 5 to 500 mg per dosage form in a unit dose; especially in the form of injection, it is preferred that the aforesaid antibody is contained in about 5 to 100 mg and in about 10 to 250 mg for the other dosage forms.

[0060] Single and Combination Therapies.

[0061] The antibodies and antibody fragments of the invention are useful for treating diseases and disorders which are improved, inhibited or ameliorated by reducing IL-4 activity. These disorders include those characterized by abnormal or excess expression of IL-4, or by an abnormal host response to IL-4 production. IL-4 related disorders which are treated by the antibodies or antibody fragments of the include, for example, arthritis (including septic arthritis), herpetiformis, chronic idiopathic urticaria, scleroderma, hypertrophic scarring, Whipple's Disease, benign prostate hyperplasia, pulmonary disorders such as asthma (mild, moderate or severe), inflammatory disorders such as inflammatory bowel disease, allergic reactions, Kawasaki disease, sickle cell disease, Churg-Strauss syndrome, Grave's disease, pre-eclampsia, Sjogren's syndrome, autoimmune lymphoproliferative syndrome, autoimmune hemolytic anemia, Barrett's esophagus, autoimmune uveitis, tuberculosis, atopic dermatatis, ulcerative colitis, fibrosis, and nephrosis (see U.S. Pat. No. 7,186,809, herein specifically incorporated by reference).

[0062] The invention encompasses combination therapies in which the anti-IL-4R antibody or antibody fragment is administered in combination with a second therapeutic agent. Co-administration and combination therapy are not limited to simultaneous administration, but include treatment regimens in which an anti-IL-4R antibody or antibody fragment is administered at least once during a course of treatment that involves administering at least one other therapeutic agent to the patient. A second therapeutic agent may be another IL-4 antagonist, such as another antibody/antibody fragment, or a soluble cytokine receptor, an IgE antagonist, an anti-asthma medication (corticosteroids, non-steroidal agents, beta agonists, leukotriene antagonists, xanthines, fluticasone, salmeterol, albuterol) which may be delivered by inhalation or other appropriate means. In a specific embodiment, the anti-IL-4R antibody or antibody fragment of the invention may be administered with an IL-1 antagonist, such as rilonacept, or an IL-13 antagonist. The second agent may include one or more leukotriene receptor antagonists to treat disorders such as allergic inflammatory diseases, e.g., asthma and allergies. Examples of leukotriene receptor antagonists include but are not limited to montelukast, pranlukast, and zafirlukast. The second agent may include a cytokine inhibitor such as one or more of a TNF (etanercept, ENBREL.TM.), IL-9, IL-5 or IL-17 antagonist.

Examples

[0063] The following examples are put forth so as to provide those of ordinary skill in the art with a complete disclosure and description of how to make and use the methods and compositions of the invention, and are not intended to limit the scope of what the inventors regard as their invention. Efforts have been made to ensure accuracy with respect to numbers used (e.g., amounts, temperature, etc.) but some experimental errors and deviations should be accounted for. Unless indicated otherwise, parts are parts by weight, molecular weight is average molecular weight, temperature is in degrees Centigrade, and pressure is at or near atmospheric.

Example 1. Generation of Human Antibodies to Human IL-4 Receptor

[0064] VELOCIMMUNE.TM. mice (Regeneron Pharmaceuticals, Inc.; U.S. Pat. No. 6,596,541) were immunized with human IL-4R (hIL-4R, SEQ ID NO:274) or a combination of hIL-4R and monkey (Macaca fascicularis) IL-4R (mfIL-4R, SEQ ID NO:275) protein or DNA. To obtain optimal immune response, animals were subsequently boosted every 3-4 weeks and bleeds obtained 10 days after each boost for assessment of progression of anti-antigen response.

[0065] When the mice attained maximum immune response, antibody-expressing B cells were harvested and fused with mouse myeloma cells to form hybridomas. Alternatively, antigen-specific antibodies may be isolated directly from the B cells without fusion to myeloma cells, as described in U.S. Patent Publication 2007/0280945A1, herein specifically incorporated by reference in its entirety. Stable recombinant antibody-expressing CHO cell lines were established from the isolated proper recombinants. Functionally desirable monoclonal antibodies were selected by screening conditioned media of the hybridomas or transfected cells for specificity, antigen-binding affinity, and potency in blocking hIL-4 binding to hIL-4R (described below).

Example 2. Antigen Binding Affinity Determination

[0066] Binding affinity (K.sub.D) of selected antibodies with respect to hIL-4R at either 25.degree. C. or 37.degree. C. was determined using a real-time biosensor surface plasmon resonance assay (BIACORE.TM. 2000). Briefly, antibody was captured on a goat anti-hFc polyclonal antibody surface created through direct coupling to a BIACORE.TM. chip to form a captured antibody surface. Various concentrations (ranging from 50 nM to 12.5 nM) of monomeric hIL-4R (R&D Systems) or dimeric hIL-4R-mFc were injected over the captured antibody surface at 10 .mu.l/min for 2.5 min at either 25.degree. C. or 37.degree. C. Binding of antigen to antibody and dissociation of the bound complex, were monitored in real time. Equilibrium dissociation constants (K.sub.D) and dissociation rate constants were ascertained by performing kinetic analysis using BIA evaluation software. BIA evaluation software was also used to calculate the half-life of antigen/antibody complex dissociation (T.sub.1/2). Results are shown in Table 1. NB: No antibody-antigen binding was observed under the experimental condition. Control: a fully human anti-IL-4R antibody (U.S. Pat. No. 7,186,809; SEQ ID NOs:10 and 12).

TABLE-US-00001 TABLE 1 25.degree. C. 37.degree. C. Monomeric Dimeric Monomeric Dimeric Antibody K.sub.D (pM) T.sub.1/2 (min) K.sub.D (pM) T.sub.1/2 (min) K.sub.D (pM) T.sub.1/2 (min) K.sub.D (pM) T.sub.1/2 (min) Control 1100 18 94 186 3970 4 114 158 H4H083P 48 361 28 245 183 87 38.1 163 H4H094P NB -- NB -- NB -- NB -- H4H095P 274 131 302 156 437 49 314 116 H4H098P 94.1 243 67.6 237 157 129 38.8 158 H4H099P NB -- NB -- NB -- NB --

[0067] Binding affinity (K.sub.D) of selected antibodies with respect to monkey (Macaca fascicularis) IL-4R (mfIL-4R) at either 25.degree. C. or 37.degree. C. was also determined using a real-time biosensor surface plasmon resonance assay described above with various concentrations (ranging from 100 nM to 25 nM) of monomeric mfIL-4R-myc-myc-his (mfIL-4R-mmh) or dimeric mfIL-4R-mFc. Only antibody H4H098P was able to bind both monomeric and dimeric mfIL-4R at 25.degree. C. with K.sub.D of 552 nM and 9.08 nM, respectively. In addition, antibody H4H098P also binds to dimeric mfIL-4R at 37.degree. C. with a K.sub.D of 24.3 nM. H4H083P had very weak binding to dimeric mfIL-4R.

[0068] Antibody-antigen binding affinity was also assessed using an ELISA-based solution competition assay. Briefly, a 96-well MAXISORP.TM. plate was first coated with 5 .mu.g/ml avidin overnight followed by BSA blocking for 1 hr. The avidin-coated plate was then incubated with 250 ng/ml biotin-hIL4 for 2 hr. The plate was used to measure either free hIL-4R-mFc (dimeric hIL-4R) or free hIL-4R-myc-myc-his (hIL4R-mmh, monomeric hIL4R) in the antibody titration sample solutions. To make the antibody titration sample, a constant amount either 25 pM of hIL-4R-mFc or 200 pM of hIL-4R-mmh was premixed with varied amounts of antibody, ranging from 0 to about 10 nM in serial dilutions, followed by 1 hr incubation at room temperature to allow antibody-antigen-binding to reach equilibrium. The equilibrated sample solutions were then transferred to the hIL-4 coated plates for measurement of either free hIL-4R-mFc or free hIL-4R-mmh. After 1 hr binding, the plate was washed and bound hIL-4R-mFc was detected using either an HRP-conjugated mouse anti-mFc polyclonal antibody or an HRP-conjugated goat anti-myc polyclonal antibodies. IC.sub.50 values were determined (Table 2).

TABLE-US-00002 TABLE 2 IC.sub.50 (pM) Antibody 25 pM hIL-4R-mFc 200 pM hIL-4R-mmh Control 8.2 87 H4H083P 9.6 80 H4H094P >10,000 >10,000 H4H095P 40 90 H4H098P 8.8 74 H4H099P >10,000 >10,000

[0069] The ELISA-based solution competition assay was also used to determine the cross reactivity of the antibodies to monkey IL-4R. Antibody H4H098P exhibits an IC.sub.50 for mfIL-4R-mFc of 300 pM and an IC.sub.50 for mfIL-4R-mmh of 20 nM.

Example 3. Neutralization of Biological Effect of hIL-4 and hIL-13 In Vitro

[0070] A bioassay was developed to determine the purified anti-hIL-4R antibodies to neutralize hIL-4R-mediated cellular function in vitro using an engineered HK293 cell line that contains human STAT6 and a STAT6 luciferase reporter. Inhibition of hIL-4R-inducible luciferase activity was determined as follows: Cells were seeded onto 96-well plates at 1.times.10.sup.4 cells/well in media and incubated overnight at 37.degree. C., 5% CO.sub.2. Antibody proteins ranging from 0 to 20 nM in serial dilutions were added to the cells along with either 10 pM hIL-4 or 40 pM of hIL-13. Cells were then incubated at 37.degree. C., 5% CO.sub.2 for 6 hrs. The extent of cellular response was measured in a luciferase assay (Promega Biotech). Results are shown in Table 3. NB: Luciferase activity was not blocked under the experimental condition described above. In addition, H4H098P was able to block mfIL-4R-mediated cellular function in the presence of 360 fM mfIL-4 with an IC.sub.50 of 150 nM.

TABLE-US-00003 TABLE 3 IC.sub.50 (pM) Antibody 10 pM hIL-4 40 pM hIL-13 Control 47 38 H4H083P 25 19 H4H094P NB NB H4H095P 98 86 H4H098P 27 25 H4H099P NB 11,000

Sequence CWU 1

1

2751351DNAArtificial SequenceSynthetic 1caggtgcagc tggtggagtc tgggggaggc gtggtccagc ctgggaggtc cctgagactc 60tcctgtgcag cctctggatt caccttccgc tcttatggca tgcactgggt ccgccaggct 120ccaggcaagg ggctggagtg ggtggcggtc atatcatatg atggaagtaa taaatattat 180atagactccg tgaagggccg attcaccatc tccagagaca attccaagaa cacgctgaat 240ctgcaaatga acagcctgag acttgaggac acggctgtat attactgtgc gaaagagggg 300agggggggat ttgactactg gggccaggga atcccggtca ccgtctcctc a 3512117PRTArtificial SequenceSynthetic 2Gln Val Gln Leu Val Glu Ser Gly Gly Gly Val Val Gln Pro Gly Arg1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Arg Ser Tyr 20 25 30 Gly Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Val Ile Ser Tyr Asp Gly Ser Asn Lys Tyr Tyr Ile Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Asn65 70 75 80 Leu Gln Met Asn Ser Leu Arg Leu Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Glu Gly Arg Gly Gly Phe Asp Tyr Trp Gly Gln Gly Ile Pro 100 105 110 Val Thr Val Ser Ser 115 324DNAArtificial SequenceSynthetic 3ggattcacct tccgctctta tggc 2448PRTArtificial SequenceSynthetic 4Gly Phe Thr Phe Arg Ser Tyr Gly1 5 524DNAArtificial SequenceSynthetic 5atatcatatg atggaagtaa taaa 2468PRTArtificial SequenceSynthetic 6Ile Ser Tyr Asp Gly Ser Asn Lys1 5 730DNAArtificial SequenceSynthetic 7gcgaaagagg ggaggggggg atttgactac 30810PRTArtificial SequenceSynthetic 8Ala Lys Glu Gly Arg Gly Gly Phe Asp Tyr1 5 10 9324DNAArtificial SequenceSynthetic 9gacatccaga tgacccagtc tccatcctca ctgtctgcat ctgtaggaga cagagtcacc 60atcacttgtc gggcgagtca ggtcataaac aattatttag cctggtttca gcagaaacca 120gggaaagtcc ctaagtccct gatccatgct gcatccagtt tacaaagtgg ggtcccatca 180aagttcagcg gcagtggatc tgggacagat ttcactctca ccatcagcag cctgcagcct 240gaagattttg caacttatta ctgccaacag tataatagtc acccgtggac gttcggccaa 300gggaccaagg tggaaatcaa acga 32410108PRTArtificial SequenceSynthetic 10Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Val Ile Asn Asn Tyr 20 25 30 Leu Ala Trp Phe Gln Gln Lys Pro Gly Lys Val Pro Lys Ser Leu Ile 35 40 45 His Ala Ala Ser Ser Leu Gln Ser Gly Val Pro Ser Lys Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Asn Ser His Pro Trp 85 90 95 Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Arg 100 105 1118DNAArtificial SequenceSynthetic 11caggtcataa acaattat 18126PRTArtificial SequenceSynthetic 12Gln Val Ile Asn Asn Tyr1 5 139DNAArtificial SequenceSynthetic 13gctgcatcc 9143PRTArtificial SequenceSynthetic 14Ala Ala Ser1 1527DNAArtificial SequenceSynthetic 15caacagtata atagtcaccc gtggacg 27169PRTArtificial SequenceSynthetic 16Gln Gln Tyr Asn Ser His Pro Trp Thr1 5 17351DNAArtificial SequenceSynthetic 17caggtgcagc tggtggagtc tgggggaggc gtggtccagc ctgggaggtc cctgagactc 60tcctgtgcag cctctggatt caccttccgc tcttatggca tgcactgggt ccgccaggct 120ccaggcaagg ggctggagtg ggtggcggtc atatcatatg atggaagtaa taaatattat 180atagactccg tgaagggccg attcaccatc tccagagaca attccaagaa cacgctgaat 240ctgcaaatga acagcctgag acttgaggac acggctgtat attactgtgc gaaagagggg 300agggggggat ttgactactg gggccaggga accctggtca ccgtctcctc a 35118117PRTArtificial SequenceSynthetic 18Gln Val Gln Leu Val Glu Ser Gly Gly Gly Val Val Gln Pro Gly Arg1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Arg Ser Tyr 20 25 30 Gly Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Val Ile Ser Tyr Asp Gly Ser Asn Lys Tyr Tyr Ile Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Asn65 70 75 80 Leu Gln Met Asn Ser Leu Arg Leu Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Glu Gly Arg Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr Leu 100 105 110 Val Thr Val Ser Ser 115 19321DNAArtificial SequenceSynthetic 19gacatccaga tgacccagtc tccatcctca ctgtctgcat ctgtaggaga cagagtcacc 60atcacttgtc gggcgagtca ggtcataaac aattatttag cctggtttca gcagaaacca 120gggaaagtcc ctaagtccct gatccatgct gcatccagtt tacaaagtgg ggtcccatca 180aagttcagcg gcagtggatc tgggacagat ttcactctca ccatcagcag cctgcagcct 240gaagattttg caacttatta ctgccaacag tataatagtc acccgtggac gttcggccaa 300gggaccaagg tggaaatcaa a 32120107PRTArtificial SequenceSynthetic 20Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Val Ile Asn Asn Tyr 20 25 30 Leu Ala Trp Phe Gln Gln Lys Pro Gly Lys Val Pro Lys Ser Leu Ile 35 40 45 His Ala Ala Ser Ser Leu Gln Ser Gly Val Pro Ser Lys Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Asn Ser His Pro Trp 85 90 95 Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys 100 105 21351DNAArtificial SequenceSynthetic 21caggtgcagc tggtggagtc tgggggaggc gtggtccagc ctgggaggtc cctgagactc 60tcctgtgcag cctctggatt caccttccgc tcttatggca tgcactgggt ccgccaggct 120ccaggcaagg ggctggagtg ggtggcagtt atatcatatg atggaagtaa taaatactat 180gcagactccg tgaagggccg attcaccatc tccagagaca attccaagaa cacgctgtat 240ctgcaaatga acagcctgag agctgaggac acggctgtgt attactgtgc gaaagagggg 300agggggggat ttgactactg gggccaggga accctggtca ccgtctcctc a 35122117PRTArtificial SequenceSynthetic 22Gln Val Gln Leu Val Glu Ser Gly Gly Gly Val Val Gln Pro Gly Arg1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Arg Ser Tyr 20 25 30 Gly Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Val Ile Ser Tyr Asp Gly Ser Asn Lys Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Glu Gly Arg Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr Leu 100 105 110 Val Thr Val Ser Ser 115 23322DNAArtificial SequenceSynthetic 23gacatccaga tgacccagtc tccatcctca ctgtctgcat ctgtaggaga cagagtcacc 60atcacttgtc gggcgagtca ggtcataaac aattatttag cctggtttca gcagaaacca 120gggaaagccc ctaagtccct gatctatgct gcatccagtt tgcaaagtgg ggtcccatca 180aggttcagcg gcagtggatc tgggacagat ttcactctca ccatcagcag cctgcagcct 240gaagattttg caacttatta ctgccaacag tataatagtc acccgtggac gttcggccaa 300gggaccaagg tggaaatcaa ac 32224107PRTArtificial SequenceSynthetic 24Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Val Ile Asn Asn Tyr 20 25 30 Leu Ala Trp Phe Gln Gln Lys Pro Gly Lys Ala Pro Lys Ser Leu Ile 35 40 45 Tyr Ala Ala Ser Ser Leu Gln Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Asn Ser His Pro Trp 85 90 95 Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys 100 105 25351DNAArtificial SequenceSynthetic 25caggtgcagc tggtggagtc tgggggaggc gtggtccagc ctgggaggtc cctgagactc 60tcctgtgcag cctctggatt caccttcaga agctatggca tacactgggt ccgccaggct 120ccaggcaagg ggctggagtg ggtggcagtt atatcatatg atggaagtaa taaatactat 180gcagactccg tgaagggccg attcaccatc tccagagaca attccaagaa cacactgtat 240ctgcaaatga acagcctgat aactgaggac acggctgtgt attattgtgt gaaagagggg 300aggggggggt ttgactactg gggccaggga accacggtca ccgtctcctc a 35126117PRTArtificial SequenceSynthetic 26Gln Val Gln Leu Val Glu Ser Gly Gly Gly Val Val Gln Pro Gly Arg1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Arg Ser Tyr 20 25 30 Gly Ile His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Val Ile Ser Tyr Asp Gly Ser Asn Lys Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Ile Thr Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Val Lys Glu Gly Arg Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr Thr 100 105 110 Val Thr Val Ser Ser 115 2724DNAArtificial SequenceSynthetic 27ggattcacct tcagaagcta tggc 24288PRTArtificial SequenceSynthetic 28Gly Phe Thr Phe Arg Ser Tyr Gly1 5 2924DNAArtificial SequenceSynthetic 29atatcatatg atggaagtaa taaa 24308PRTArtificial SequenceSynthetic 30Ile Ser Tyr Asp Gly Ser Asn Lys1 5 3130DNAArtificial SequenceSynthetic 31gtgaaagagg ggaggggggg gtttgactac 303210PRTArtificial SequenceSynthetic 32Val Lys Glu Gly Arg Gly Gly Phe Asp Tyr1 5 10 33324DNAArtificial SequenceSynthetic 33gacatccaga tgacccagtc tccatcctca ctgtctgcat ctgtaggaga cagagtcacc 60atcacttgtc gggcgagtca ggtcattaat aattatttag cctggtttca gcagaaacca 120gggaaagtcc ctaagtccct gatccatgct gcatccagtt tgcaaagagg ggtcccatca 180aagttcagcg gcagtggatc tgggacagat ttcactctca ccatcaacag cctgcagcct 240gaagattttg caacttatta ctgccaacaa tataatagtt acccgtggac gttcggccaa 300gggaccaagg tggaaatcaa acga 32434108PRTArtificial SequenceSynthetic 34Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Val Ile Asn Asn Tyr 20 25 30 Leu Ala Trp Phe Gln Gln Lys Pro Gly Lys Val Pro Lys Ser Leu Ile 35 40 45 His Ala Ala Ser Ser Leu Gln Arg Gly Val Pro Ser Lys Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Asn Ser Leu Gln Pro65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Asn Ser Tyr Pro Trp 85 90 95 Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Arg 100 105 3518DNAArtificial SequenceSynthetic 35caggtcatta ataattat 18366PRTArtificial SequenceSynthetic 36Gln Val Ile Asn Asn Tyr1 5 379DNAArtificial SequenceSynthetic 37gctgcatcc 9383PRTArtificial SequenceSynthetic 38Ala Ala Ser1 3927DNAArtificial SequenceSynthetic 39caacaatata atagttaccc gtggacg 27409PRTArtificial SequenceSynthetic 40Gln Gln Tyr Asn Ser Tyr Pro Trp Thr1 5 41351DNAArtificial SequenceSynthetic 41caggtgcagc tggtggagtc tgggggaggc gtggtccagc ctgggaggtc cctgagactc 60tcctgtgcag cctctggatt caccttcaga agctatggca tacactgggt ccgccaggct 120ccaggcaagg ggctggagtg ggtggcagtt atatcatatg atggaagtaa taaatactat 180gcagactccg tgaagggccg attcaccatc tccagagaca attccaagaa cacactgtat 240ctgcaaatga acagcctgat aactgaggac acggctgtgt attattgtgt gaaagagggg 300aggggggggt ttgactactg gggccaggga accctggtca ccgtctcctc a 35142117PRTArtificial SequenceSynthetic 42Gln Val Gln Leu Val Glu Ser Gly Gly Gly Val Val Gln Pro Gly Arg1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Arg Ser Tyr 20 25 30 Gly Ile His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Val Ile Ser Tyr Asp Gly Ser Asn Lys Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Ile Thr Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Val Lys Glu Gly Arg Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr Leu 100 105 110 Val Thr Val Ser Ser 115 43321DNAArtificial SequenceSynthetic 43gacatccaga tgacccagtc tccatcctca ctgtctgcat ctgtaggaga cagagtcacc 60atcacttgtc gggcgagtca ggtcattaat aattatttag cctggtttca gcagaaacca 120gggaaagtcc ctaagtccct gatccatgct gcatccagtt tgcaaagagg ggtcccatca 180aagttcagcg gcagtggatc tgggacagat ttcactctca ccatcaacag cctgcagcct 240gaagattttg caacttatta ctgccaacaa tataatagtt acccgtggac gttcggccaa 300gggaccaagg tggaaatcaa a 32144107PRTArtificial SequenceSynthetic 44Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Val Ile Asn Asn Tyr 20 25 30 Leu Ala Trp Phe Gln Gln Lys Pro Gly Lys Val Pro Lys Ser Leu Ile 35 40 45 His Ala Ala Ser Ser Leu Gln Arg Gly Val Pro Ser Lys Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Asn Ser Leu Gln Pro65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Asn Ser Tyr Pro Trp 85 90 95 Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys 100 105 45351DNAArtificial SequenceSynthetic 45caggtgcagc tggtggagtc tgggggaggc gtggtccagc ctgggaggtc cctgagactc 60tcctgtgcag cctctggatt caccttcaga agctatggca tgcactgggt ccgccaggct 120ccaggcaagg ggctggagtg ggtggcagtt atatcatatg atggaagtaa taaatactat 180gcagactccg tgaagggccg attcaccatc tccagagaca attccaagaa cacgctgtat 240ctgcaaatga acagcctgag agctgaggac acggctgtgt attactgtgt gaaagagggg 300aggggggggt ttgactactg gggccaggga accctggtca ccgtctcctc a 35146117PRTArtificial SequenceSynthetic 46Gln Val Gln Leu Val Glu Ser Gly Gly Gly Val Val Gln Pro Gly Arg1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Arg Ser Tyr 20 25 30 Gly Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Val Ile Ser Tyr Asp Gly Ser Asn Lys Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Val Lys Glu Gly Arg Gly Gly Phe Asp Tyr Trp Gly Gln Gly Thr Leu 100 105 110 Val Thr Val Ser Ser 115 47322DNAArtificial SequenceSynthetic 47gacatccaga tgacccagtc tccatcctca ctgtctgcat ctgtaggaga cagagtcacc 60atcacttgtc gggcgagtca ggtcattaat aattatttag

cctggtttca gcagaaacca 120gggaaagccc ctaagtccct gatctatgct gcatccagtt tgcaaagtgg ggtcccatca 180aggttcagcg gcagtggatc tgggacagat ttcactctca ccatcagcag cctgcagcct 240gaagattttg caacttatta ctgccaacaa tataatagtt acccgtggac gttcggccaa 300gggaccaagg tggaaatcaa ac 32248107PRTArtificial SequenceSynthetic 48Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Val Ile Asn Asn Tyr 20 25 30 Leu Ala Trp Phe Gln Gln Lys Pro Gly Lys Ala Pro Lys Ser Leu Ile 35 40 45 Tyr Ala Ala Ser Ser Leu Gln Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Asn Ser Tyr Pro Trp 85 90 95 Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys 100 105 49375DNAArtificial SequenceSynthetic 49caggtgcagc tggtggagtc tgggggaggc ttggaacagc cgggggggtc cttgagactc 60tcctgtgcag gctctggatt cacgtttaga gactatgcca tgacctgggt ccgccaggct 120ccagggaagg ggctggagtg ggtcgcatcg attagtggtt ccggtggtaa cacatacttc 180gcagactccg tgaagggccg gttcaccatc tccagagaca attccaagaa cacgctgtat 240ctgcaaatga acagcctgag agccgaggac acggccgtat attactgtgc gaaagatcga 300ctctctataa caattcgccc acgctattat ggtttggacg tctggggcca agggtccacg 360gtcaccgtct cctca 37550125PRTArtificial SequenceSynthetic 50Gln Val Gln Leu Val Glu Ser Gly Gly Gly Leu Glu Gln Pro Gly Gly1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Gly Ser Gly Phe Thr Phe Arg Asp Tyr 20 25 30 Ala Met Thr Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Ser Ile Ser Gly Ser Gly Gly Asn Thr Tyr Phe Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Asp Arg Leu Ser Ile Thr Ile Arg Pro Arg Tyr Tyr Gly Leu 100 105 110 Asp Val Trp Gly Gln Gly Ser Thr Val Thr Val Ser Ser 115 120 125 5124DNAArtificial SequenceSynthetic 51ggattcacgt ttagagacta tgcc 24528PRTArtificial SequenceSynthetic 52Gly Phe Thr Phe Arg Asp Tyr Ala1 5 5324DNAArtificial SequenceSynthetic 53attagtggtt ccggtggtaa caca 24548PRTArtificial SequenceSynthetic 54Ile Ser Gly Ser Gly Gly Asn Thr1 5 5554DNAArtificial SequenceSynthetic 55gcgaaagatc gactctctat aacaattcgc ccacgctatt atggtttgga cgtc 545618PRTArtificial SequenceSynthetic 56Ala Lys Asp Arg Leu Ser Ile Thr Ile Arg Pro Arg Tyr Tyr Gly Leu1 5 10 15 Asp Val57324DNAArtificial SequenceSynthetic 57gacatccaga tgacccagtc tccatcctca ctgtctgcat ctgttggaga cagagtcacc 60atcacttgtc gggcgagtca ggccattaac aatcatttag cctggtttca gcagaaacca 120gggaaagccc ctaagtccct gatctttgct gtatccagtt tgcaaagtgg ggtcccatca 180aagttcagcg gcagtggatc tgggacagac ttcactctca ccatcagcag cctgcagcct 240gaagattttg caacttatta ctgccaacag tataatagtt acccgtggac gttcggccaa 300gggaccaagg tggaaatcaa acga 32458108PRTArtificial SequenceSynthetic 58Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Ala Ile Asn Asn His 20 25 30 Leu Ala Trp Phe Gln Gln Lys Pro Gly Lys Ala Pro Lys Ser Leu Ile 35 40 45 Phe Ala Val Ser Ser Leu Gln Ser Gly Val Pro Ser Lys Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Asn Ser Tyr Pro Trp 85 90 95 Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Arg 100 105 5918DNAArtificial SequenceSynthetic 59caggccatta acaatcat 18606PRTArtificial SequenceSynthetic 60Gln Ala Ile Asn Asn His1 5 619DNAArtificial SequenceSynthetic 61gctgtatcc 9623PRTArtificial SequenceSynthetic 62Ala Val Ser1 6327DNAArtificial SequenceSynthetic 63caacagtata atagttaccc gtggacg 27649PRTArtificial SequenceSynthetic 64Gln Gln Tyr Asn Ser Tyr Pro Trp Thr1 5 65372DNAArtificial SequenceSynthetic 65gaggtgcagc tggtggagtc tgggggaggc ttggaacagc cgggggggtc cttgagactc 60tcctgtgcag gctctggatt cacgtttaga gactatgcca tgacctgggt ccgccaggct 120ccagggaagg ggctggagtg ggtcgcatcg attagtggtt ccggtggtaa cacatacttc 180gcagactccg tgaagggccg gttcaccatc tccagagaca attccaagaa cacgctgtat 240ctgcaaatga acagcctgag agccgaggac acggccgtat attactgtgc gaaagatcga 300ctctctataa caattcgccc acgctattat ggtttggacg tctggggcca agggaccacg 360gtcaccgtct cc 37266124PRTArtificial SequenceSynthetic 66Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Glu Gln Pro Gly Gly1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Gly Ser Gly Phe Thr Phe Arg Asp Tyr 20 25 30 Ala Met Thr Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Ser Ile Ser Gly Ser Gly Gly Asn Thr Tyr Phe Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Asp Arg Leu Ser Ile Thr Ile Arg Pro Arg Tyr Tyr Gly Leu 100 105 110 Asp Val Trp Gly Gln Gly Thr Thr Val Thr Val Ser 115 120 67321DNAArtificial SequenceSynthetic 67gacatccaga tgacccagtc tccatcctca ctgtctgcat ctgttggaga cagagtcacc 60atcacttgtc gggcgagtca ggccattaac aatcatttag cctggtttca gcagaaacca 120gggaaagccc ctaagtccct gatctttgct gtatccagtt tgcaaagtgg ggtcccatca 180aagttcagcg gcagtggatc tgggacagac ttcactctca ccatcagcag cctgcagcct 240gaagattttg caacttatta ctgccaacag tataatagtt acccgtggac gttcggccaa 300gggaccaagg tggaaatcaa a 32168107PRTArtificial SequenceSynthetic 68Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Ala Ile Asn Asn His 20 25 30 Leu Ala Trp Phe Gln Gln Lys Pro Gly Lys Ala Pro Lys Ser Leu Ile 35 40 45 Phe Ala Val Ser Ser Leu Gln Ser Gly Val Pro Ser Lys Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Asn Ser Tyr Pro Trp 85 90 95 Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys 100 105 69373DNAArtificial SequenceSynthetic 69gaggtgcagc tggtggagtc tgggggaggc ttggtacagc ctggggggtc cctgagactc 60tcctgtgcag cctctggatt cacgtttaga gactatgcca tgagctgggt ccgccaggct 120ccagggaagg ggctggagtg ggtctcagct attagtggtt ccggtggtaa cacatactac 180gcagactccg tgaagggccg gttcaccatc tccagagaca attccaagaa cacgctgtat 240ctgcaaatga acagcctgag agccgaggac acggccgtat attactgtgc gaaagatcga 300ctctctataa caattcgccc acgctattat ggtttggacg tctggggcca agggaccacg 360gtcaccgtct cct 37370124PRTArtificial SequenceSynthetic 70Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Arg Asp Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Gly Ser Gly Gly Asn Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Asp Arg Leu Ser Ile Thr Ile Arg Pro Arg Tyr Tyr Gly Leu 100 105 110 Asp Val Trp Gly Gln Gly Thr Thr Val Thr Val Ser 115 120 71322DNAArtificial SequenceSynthetic 71gacatccaga tgacccagtc tccatcctca ctgtctgcat ctgtaggaga cagagtcacc 60atcacttgtc gggcgagtca ggccattaac aatcatttag cctggtttca gcagaaacca 120gggaaagccc ctaagtccct gatctatgct gtatccagtt tgcaaagtgg ggtcccatca 180aggttcagcg gcagtggatc tgggacagat ttcactctca ccatcagcag cctgcagcct 240gaagattttg caacttatta ctgccaacag tataatagtt acccgtggac gttcggccaa 300gggaccaagg tggaaatcaa ac 32272107PRTArtificial SequenceSynthetic 72Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Ala Ile Asn Asn His 20 25 30 Leu Ala Trp Phe Gln Gln Lys Pro Gly Lys Ala Pro Lys Ser Leu Ile 35 40 45 Tyr Ala Val Ser Ser Leu Gln Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Tyr Asn Ser Tyr Pro Trp 85 90 95 Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys 100 105 73375DNAArtificial SequenceSynthetic 73caggtgcagc tggtggagtc tgggggaggc ttggaacagc cgggggggtc cttgagactc 60tcctgtgcag gctctggatt cacgtttaga gactatgcca tgacctgggt ccgccaggct 120ccagggaagg ggctggagtg ggtcgcatcg attagtggtt ccggtggtaa cacatacttc 180gcagactccg tgaagggccg gttcaccatc tccagagaca attccaagaa cacgctgtat 240ctgcaaatga acagcctgag agccgaggac acggccgtat attactgtgc gaaagatcga 300ctctctataa caattcgccc acgctattat ggtttggacg tctggggcca agggtccacg 360gtcaccgtct cctca 37574125PRTArtificial SequenceSynthetic 74Gln Val Gln Leu Val Glu Ser Gly Gly Gly Leu Glu Gln Pro Gly Gly1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Gly Ser Gly Phe Thr Phe Arg Asp Tyr 20 25 30 Ala Met Thr Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Ser Ile Ser Gly Ser Gly Gly Asn Thr Tyr Phe Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Asp Arg Leu Ser Ile Thr Ile Arg Pro Arg Tyr Tyr Gly Leu 100 105 110 Asp Val Trp Gly Gln Gly Ser Thr Val Thr Val Ser Ser 115 120 125 7524DNAArtificial SequenceSynthetic 75ggattcacgt ttagagacta tgcc 24768PRTArtificial SequenceSynthetic 76Gly Phe Thr Phe Arg Asp Tyr Ala1 5 7724DNAArtificial SequenceSynthetic 77attagtggtt ccggtggtaa caca 24788PRTArtificial SequenceSynthetic 78Ile Ser Gly Ser Gly Gly Asn Thr1 5 7954DNAArtificial SequenceSynthetic 79gcgaaagatc gactctctat aacaattcgc ccacgctatt atggtttgga cgtc 548018PRTArtificial SequenceSynthetic 80Ala Lys Asp Arg Leu Ser Ile Thr Ile Arg Pro Arg Tyr Tyr Gly Leu1 5 10 15 Asp Val81339DNAArtificial SequenceSynthetic 81gaaatagtgt tgacgcagtc tccactctcc ctgcccgtca cccctggaga gccggcctcc 60atctcctgca ggtctagtca gagcctcctg tatagtattg gatacaacta tttggattgg 120tacctgcaga agtcagggca gtctccacag ctccttatct atttgggttc taatcgggcc 180tccggggtcc ctgacaggtt cagtggcagt ggatcaggca cagattttac actgaaaatc 240agcagagtgg aggctgagga tgttgggttt tattactgca tgcaagctct acaaactccg 300tacacttttg gcccggggac caagctggag atcaaacga 33982113PRTArtificial SequenceSynthetic 82Glu Ile Val Leu Thr Gln Ser Pro Leu Ser Leu Pro Val Thr Pro Gly1 5 10 15 Glu Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Leu Leu Tyr Ser 20 25 30 Ile Gly Tyr Asn Tyr Leu Asp Trp Tyr Leu Gln Lys Ser Gly Gln Ser 35 40 45 Pro Gln Leu Leu Ile Tyr Leu Gly Ser Asn Arg Ala Ser Gly Val Pro 50 55 60 Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile65 70 75 80 Ser Arg Val Glu Ala Glu Asp Val Gly Phe Tyr Tyr Cys Met Gln Ala 85 90 95 Leu Gln Thr Pro Tyr Thr Phe Gly Pro Gly Thr Lys Leu Glu Ile Lys 100 105 110 Arg 8333DNAArtificial SequenceSynthetic 83cagagcctcc tgtatagtat tggatacaac tat 338411PRTArtificial SequenceSynthetic 84Gln Ser Leu Leu Tyr Ser Ile Gly Tyr Asn Tyr1 5 10 859DNAArtificial SequenceSynthetic 85ttgggttct 9863PRTArtificial SequenceSynthetic 86Leu Gly Ser1 8727DNAArtificial SequenceSynthetic 87atgcaagctc tacaaactcc gtacact 27889PRTArtificial SequenceSynthetic 88Met Gln Ala Leu Gln Thr Pro Tyr Thr1 5 89372DNAArtificial SequenceSynthetic 89gaggtgcagc tggtggagtc tgggggaggc ttggaacagc cgggggggtc cttgagactc 60tcctgtgcag gctctggatt cacgtttaga gactatgcca tgacctgggt ccgccaggct 120ccagggaagg ggctggagtg ggtcgcatcg attagtggtt ccggtggtaa cacatacttc 180gcagactccg tgaagggccg gttcaccatc tccagagaca attccaagaa cacgctgtat 240ctgcaaatga acagcctgag agccgaggac acggccgtat attactgtgc gaaagatcga 300ctctctataa caattcgccc acgctattat ggtttggacg tctggggcca agggaccacg 360gtcaccgtct cc 37290124PRTArtificial SequenceSynthetic 90Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Glu Gln Pro Gly Gly1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Gly Ser Gly Phe Thr Phe Arg Asp Tyr 20 25 30 Ala Met Thr Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Ser Ile Ser Gly Ser Gly Gly Asn Thr Tyr Phe Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Asp Arg Leu Ser Ile Thr Ile Arg Pro Arg Tyr Tyr Gly Leu 100 105 110 Asp Val Trp Gly Gln Gly Thr Thr Val Thr Val Ser 115 120 91336DNAArtificial SequenceSynthetic 91gatattgtga tgactcagtc tccactctcc ctgcccgtca cccctggaga gccggcctcc 60atctcctgca ggtctagtca gagcctcctg tatagtattg gatacaacta tttggattgg 120tacctgcaga agtcagggca gtctccacag ctccttatct atttgggttc taatcgggcc 180tccggggtcc ctgacaggtt cagtggcagt ggatcaggca cagattttac actgaaaatc 240agcagagtgg aggctgagga tgttgggttt tattactgca tgcaagctct acaaactccg 300tacacttttg gcccggggac caagctggag atcaaa 33692112PRTArtificial SequenceSynthetic 92Asp Ile Val Met Thr Gln Ser Pro Leu Ser Leu Pro Val Thr Pro Gly1 5 10 15 Glu Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Leu Leu Tyr Ser 20 25 30 Ile Gly Tyr Asn Tyr Leu Asp Trp Tyr Leu Gln Lys Ser Gly Gln Ser 35 40 45 Pro Gln Leu Leu Ile Tyr Leu Gly Ser Asn Arg Ala Ser Gly Val Pro 50 55 60 Asp Arg Phe Ser Gly Ser Gly Ser Gly

Thr Asp Phe Thr Leu Lys Ile65 70 75 80 Ser Arg Val Glu Ala Glu Asp Val Gly Phe Tyr Tyr Cys Met Gln Ala 85 90 95 Leu Gln Thr Pro Tyr Thr Phe Gly Pro Gly Thr Lys Leu Glu Ile Lys 100 105 110 93373DNAArtificial SequenceSynthetic 93gaggtgcagc tggtggagtc tgggggaggc ttggtacagc ctggggggtc cctgagactc 60tcctgtgcag cctctggatt cacgtttaga gactatgcca tgagctgggt ccgccaggct 120ccagggaagg ggctggagtg ggtctcagct attagtggtt ccggtggtaa cacatactac 180gcagactccg tgaagggccg gttcaccatc tccagagaca attccaagaa cacgctgtat 240ctgcaaatga acagcctgag agccgaggac acggccgtat attactgtgc gaaagatcga 300ctctctataa caattcgccc acgctattat ggtttggacg tctggggcca agggaccacg 360gtcaccgtct cct 37394124PRTArtificial SequenceSynthetic 94Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Arg Asp Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Gly Ser Gly Gly Asn Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Asp Arg Leu Ser Ile Thr Ile Arg Pro Arg Tyr Tyr Gly Leu 100 105 110 Asp Val Trp Gly Gln Gly Thr Thr Val Thr Val Ser 115 120 95337DNAArtificial SequenceSynthetic 95gatattgtga tgactcagtc tccactctcc ctgcccgtca cccctggaga gccggcctcc 60atctcctgca ggtctagtca gagcctcctg tatagtattg gatacaacta tttggattgg 120tacctgcaga agccagggca gtctccacag ctcctgatct atttgggttc taatcgggcc 180tccggggtcc ctgacaggtt cagtggcagt ggatcaggca cagattttac actgaaaatc 240agcagagtgg aggctgagga tgttggggtt tattactgca tgcaagctct acaaactccg 300tacacttttg gccaggggac caagctggag atcaaac 33796112PRTArtificial SequenceSynthetic 96Asp Ile Val Met Thr Gln Ser Pro Leu Ser Leu Pro Val Thr Pro Gly1 5 10 15 Glu Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Leu Leu Tyr Ser 20 25 30 Ile Gly Tyr Asn Tyr Leu Asp Trp Tyr Leu Gln Lys Pro Gly Gln Ser 35 40 45 Pro Gln Leu Leu Ile Tyr Leu Gly Ser Asn Arg Ala Ser Gly Val Pro 50 55 60 Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile65 70 75 80 Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Met Gln Ala 85 90 95 Leu Gln Thr Pro Tyr Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 110 97375DNAArtificial SequenceSynthetic 97caggtgcagc tggtggagtc tgagggactc ttggaacagc ctggggggtc cctgagactc 60tcctgtgcag cctctggatt caactttaga gactttgcca tgacctgggt ccgccaggct 120ccagggaagg ggctggagtg ggtctcatct attagtggta gtggtagtaa tacatactac 180gcagactccg tgaagggccg gttcaccatc tccagagaca attccaacca cacgctgtat 240ctgcgaatga acagcctgag agccgaagac acggccgtgt attactgtgc gaaagatcga 300ctctctataa caattcgccc acgctattac ggtctggacg tctggggcca agggtccacg 360gtcaccgtct cctca 37598125PRTArtificial SequenceSynthetic 98Gln Val Gln Leu Val Glu Ser Glu Gly Leu Leu Glu Gln Pro Gly Gly1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Asn Phe Arg Asp Phe 20 25 30 Ala Met Thr Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Ile Ser Gly Ser Gly Ser Asn Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Asn His Thr Leu Tyr65 70 75 80 Leu Arg Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Asp Arg Leu Ser Ile Thr Ile Arg Pro Arg Tyr Tyr Gly Leu 100 105 110 Asp Val Trp Gly Gln Gly Ser Thr Val Thr Val Ser Ser 115 120 125 9924DNAArtificial SequenceSynthetic 99ggattcaact ttagagactt tgcc 241008PRTArtificial SequenceSynthetic 100Gly Phe Asn Phe Arg Asp Phe Ala1 5 10124DNAArtificial SequenceSynthetic 101attagtggta gtggtagtaa taca 241028PRTArtificial SequenceSynthetic 102Ile Ser Gly Ser Gly Ser Asn Thr1 5 10354DNAArtificial SequenceSynthetic 103gcgaaagatc gactctctat aacaattcgc ccacgctatt acggtctgga cgtc 5410418PRTArtificial SequenceSynthetic 104Ala Lys Asp Arg Leu Ser Ile Thr Ile Arg Pro Arg Tyr Tyr Gly Leu1 5 10 15 Asp Val105324DNAArtificial SequenceSynthetic 105gacatccaga tgacccagtc tccatcctcc ctgtctgcat ctgtaggaga cagagtcacc 60atcacttgcc gggcgagtca ggacattagc aattattttg cctggtatca gcagaagcca 120gggaaagttc ctaagctcct gatctttgct gcatccactt tgcatccagg ggtcccatct 180cggttcagtg gcagtggatc tgggacagat ttcactctca ccattcgcag cctgcagcct 240gaagatgttg caacttatta ctgtcaaaaa tatgacagtg ccccgtacac ttttggccag 300gggaccaagg tggaaatcaa acga 324106108PRTArtificial SequenceSynthetic 106Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Ile Ser Asn Tyr 20 25 30 Phe Ala Trp Tyr Gln Gln Lys Pro Gly Lys Val Pro Lys Leu Leu Ile 35 40 45 Phe Ala Ala Ser Thr Leu His Pro Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Arg Ser Leu Gln Pro65 70 75 80 Glu Asp Val Ala Thr Tyr Tyr Cys Gln Lys Tyr Asp Ser Ala Pro Tyr 85 90 95 Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Arg 100 105 10718DNAArtificial SequenceSynthetic 107caggacatta gcaattat 181086PRTArtificial SequenceSynthetic 108Gln Asp Ile Ser Asn Tyr1 5 1099DNAArtificial SequenceSynthetic 109gctgcatcc 91103PRTArtificial SequenceSynthetic 110Ala Ala Ser1 11127DNAArtificial SequenceSynthetic 111caaaaatatg acagtgcccc gtacact 271129PRTArtificial SequenceSynthetic 112Gln Lys Tyr Asp Ser Ala Pro Tyr Thr1 5 113372DNAArtificial SequenceSynthetic 113gaggtgcagc tggtggagtc tgagggactc ttggaacagc ctggggggtc cctgagactc 60tcctgtgcag cctctggatt caactttaga gactttgcca tgacctgggt ccgccaggct 120ccagggaagg ggctggagtg ggtctcatct attagtggta gtggtagtaa tacatactac 180gcagactccg tgaagggccg gttcaccatc tccagagaca attccaacca cacgctgtat 240ctgcgaatga acagcctgag agccgaagac acggccgtgt attactgtgc gaaagatcga 300ctctctataa caattcgccc acgctattac ggtctggacg tctggggcca agggaccacg 360gtcaccgtct cc 372114124PRTArtificial SequenceSynthetic 114Glu Val Gln Leu Val Glu Ser Glu Gly Leu Leu Glu Gln Pro Gly Gly1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Asn Phe Arg Asp Phe 20 25 30 Ala Met Thr Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Ile Ser Gly Ser Gly Ser Asn Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Asn His Thr Leu Tyr65 70 75 80 Leu Arg Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Asp Arg Leu Ser Ile Thr Ile Arg Pro Arg Tyr Tyr Gly Leu 100 105 110 Asp Val Trp Gly Gln Gly Thr Thr Val Thr Val Ser 115 120 115321DNAArtificial SequenceSynthetic 115gacatccaga tgacccagtc tccatcctcc ctgtctgcat ctgtaggaga cagagtcacc 60atcacttgcc gggcgagtca ggacattagc aattattttg cctggtatca gcagaagcca 120gggaaagttc ctaagctcct gatctttgct gcatccactt tgcatccagg ggtcccatct 180cggttcagtg gcagtggatc tgggacagat ttcactctca ccattcgcag cctgcagcct 240gaagatgttg caacttatta ctgtcaaaaa tatgacagtg ccccgtacac ttttggccag 300gggaccaagc tggagatcaa a 321116107PRTArtificial SequenceSynthetic 116Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Ile Ser Asn Tyr 20 25 30 Phe Ala Trp Tyr Gln Gln Lys Pro Gly Lys Val Pro Lys Leu Leu Ile 35 40 45 Phe Ala Ala Ser Thr Leu His Pro Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Arg Ser Leu Gln Pro65 70 75 80 Glu Asp Val Ala Thr Tyr Tyr Cys Gln Lys Tyr Asp Ser Ala Pro Tyr 85 90 95 Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 117373DNAArtificial SequenceSynthetic 117gaggtgcagc tggtggagtc tgggggaggc ttggtacagc ctggggggtc cctgagactc 60tcctgtgcag cctctggatt caactttaga gactttgcca tgagctgggt ccgccaggct 120ccagggaagg ggctggagtg ggtctcagct attagtggta gtggtagtaa tacatactac 180gcagactccg tgaagggccg gttcaccatc tccagagaca attccaagaa cacgctgtat 240ctgcaaatga acagcctgag agccgaggac acggccgtat attactgtgc gaaagatcga 300ctctctataa caattcgccc acgctattac ggtctggacg tctggggcca agggaccacg 360gtcaccgtct cct 373118124PRTArtificial SequenceSynthetic 118Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Asn Phe Arg Asp Phe 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Gly Ser Gly Ser Asn Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Asp Arg Leu Ser Ile Thr Ile Arg Pro Arg Tyr Tyr Gly Leu 100 105 110 Asp Val Trp Gly Gln Gly Thr Thr Val Thr Val Ser 115 120 119322DNAArtificial SequenceSynthetic 119gacatccaga tgacccagtc tccatcctcc ctgtctgcat ctgtaggaga cagagtcacc 60atcacttgcc gggcgagtca ggacattagc aattatttag cctggtatca gcagaaacca 120gggaaagttc ctaagctcct gatctatgct gcatccactt tgcaatcagg ggtcccatct 180cggttcagtg gcagtggatc tgggacagat ttcactctca ccatcagcag cctgcagcct 240gaagatgttg caacttatta ctgtcaaaaa tatgacagtg ccccgtacac ttttggccag 300gggaccaagc tggagatcaa ac 322120107PRTArtificial SequenceSynthetic 120Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Ile Ser Asn Tyr 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Lys Val Pro Lys Leu Leu Ile 35 40 45 Tyr Ala Ala Ser Thr Leu Gln Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro65 70 75 80 Glu Asp Val Ala Thr Tyr Tyr Cys Gln Lys Tyr Asp Ser Ala Pro Tyr 85 90 95 Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 121357DNAArtificial SequenceSynthetic 121caggtgcagc tggtggagtc tgggggaggc ttggtacagc ctggggggtc cctgagactc 60tcctgtgtag cttctggatt cacccttaac aactttgtca tgaactgggt ccgccaggtt 120ccagggaagg gactggagtg ggtctctttt attagtgcta gtggtggtag tatatactac 180gcagactccg tgaagggccg gttcaccatc tccagagaca cttccaagaa cacattatat 240ctgcaaatga acagcctgag agccgacgac acggccgtct attactgtgc gaaatccccg 300tataactgga acccctttga ctattggggc cagggaacca cggtcaccgt ctcctca 357122119PRTArtificial SequenceSynthetic 122Gln Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly1 5 10 15 Ser Leu Arg Leu Ser Cys Val Ala Ser Gly Phe Thr Leu Asn Asn Phe 20 25 30 Val Met Asn Trp Val Arg Gln Val Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Phe Ile Ser Ala Ser Gly Gly Ser Ile Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Thr Ser Lys Asn Thr Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Ser Pro Tyr Asn Trp Asn Pro Phe Asp Tyr Trp Gly Gln Gly 100 105 110 Thr Thr Val Thr Val Ser Ser 115 12324DNAArtificial SequenceSynthetic 123ggattcaccc ttaacaactt tgtc 241248PRTArtificial SequenceSynthetic 124Gly Phe Thr Leu Asn Asn Phe Val1 5 12524DNAArtificial SequenceSynthetic 125attagtgcta gtggtggtag tata 241268PRTArtificial SequenceSynthetic 126Ile Ser Ala Ser Gly Gly Ser Ile1 5 12736DNAArtificial SequenceSynthetic 127gcgaaatccc cgtataactg gaaccccttt gactat 3612812PRTArtificial SequenceSynthetic 128Ala Lys Ser Pro Tyr Asn Trp Asn Pro Phe Asp Tyr1 5 10 129327DNAArtificial SequenceSynthetic 129gacatccagt tgacccagtc tccagccacc ctgtctgtgt ctccagggga acgagccacc 60ctctcctgca gggccagtct gagtgttagc agcaaattag cctggtacca gcagacacct 120ggccaggctc ccagactcct catctatagt gcctccaccc gggccactgg tatcccagtc 180aggttcagtg gcagtgggtc tgggacagag ttcactctca ccatcagcag cctgcagtct 240gaagattttg cggtttatta ctgtcagcag tataatcatt ggcctccgta cacttttggc 300caggggacca aggtggagat caaacga 327130109PRTArtificial SequenceSynthetic 130Asp Ile Gln Leu Thr Gln Ser Pro Ala Thr Leu Ser Val Ser Pro Gly1 5 10 15 Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Leu Ser Val Ser Ser Lys 20 25 30 Leu Ala Trp Tyr Gln Gln Thr Pro Gly Gln Ala Pro Arg Leu Leu Ile 35 40 45 Tyr Ser Ala Ser Thr Arg Ala Thr Gly Ile Pro Val Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Glu Phe Thr Leu Thr Ile Ser Ser Leu Gln Ser65 70 75 80 Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Tyr Asn His Trp Pro Pro 85 90 95 Tyr Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Arg 100 105 13118DNAArtificial SequenceSynthetic 131ctgagtgtta gcagcaaa 181326PRTArtificial SequenceSynthetic 132Leu Ser Val Ser Ser Lys1 5 1339DNAArtificial SequenceSynthetic 133agtgcctcc 91343PRTArtificial SequenceSynthetic 134Ser Ala Ser1 13530DNAArtificial SequenceSynthetic 135cagcagtata atcattggcc tccgtacact 3013610PRTArtificial SequenceSynthetic 136Gln Gln Tyr Asn His Trp Pro Pro Tyr Thr1 5 10 137357DNAArtificial SequenceSynthetic 137gaggtgcagc tggtggagtc tgggggaggc ttggtacagc ctggggggtc cctgagactc 60tcctgtgtag cttctggatt cacccttaac aactttgtca tgaactgggt ccgccaggtt 120ccagggaagg gactggagtg ggtctctttt attagtgcta gtggtggtag tatatactac 180gcagactccg tgaagggccg gttcaccatc tccagagaca cttccaagaa cacattatat 240ctgcaaatga acagcctgag agccgacgac acggccgtct attactgtgc gaaatccccg 300tataactgga acccctttga ctattggggc cagggaaccc tggtcaccgt ctcctca 357138119PRTArtificial SequenceSynthetic 138Glu

Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly1 5 10 15 Ser Leu Arg Leu Ser Cys Val Ala Ser Gly Phe Thr Leu Asn Asn Phe 20 25 30 Val Met Asn Trp Val Arg Gln Val Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Phe Ile Ser Ala Ser Gly Gly Ser Ile Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Thr Ser Lys Asn Thr Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Ser Pro Tyr Asn Trp Asn Pro Phe Asp Tyr Trp Gly Gln Gly 100 105 110 Thr Leu Val Thr Val Ser Ser 115 139324DNAArtificial SequenceSynthetic 139gaaatagtga tgacgcagtc tccagccacc ctgtctgtgt ctccagggga acgagccacc 60ctctcctgca gggccagtct gagtgttagc agcaaattag cctggtacca gcagacacct 120ggccaggctc ccagactcct catctatagt gcctccaccc gggccactgg tatcccagtc 180aggttcagtg gcagtgggtc tgggacagag ttcactctca ccatcagcag cctgcagtct 240gaagattttg cggtttatta ctgtcagcag tataatcatt ggcctccgta cacttttggc 300caggggacca agctggagat caaa 324140108PRTArtificial SequenceSynthetic 140Glu Ile Val Met Thr Gln Ser Pro Ala Thr Leu Ser Val Ser Pro Gly1 5 10 15 Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Leu Ser Val Ser Ser Lys 20 25 30 Leu Ala Trp Tyr Gln Gln Thr Pro Gly Gln Ala Pro Arg Leu Leu Ile 35 40 45 Tyr Ser Ala Ser Thr Arg Ala Thr Gly Ile Pro Val Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Glu Phe Thr Leu Thr Ile Ser Ser Leu Gln Ser65 70 75 80 Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Tyr Asn His Trp Pro Pro 85 90 95 Tyr Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 141357DNAArtificial SequenceSynthetic 141gaggtgcagc tggtggagtc tgggggaggc ttggtacagc ctggggggtc cctgagactc 60tcctgtgcag cctctggatt cacccttaac aactttgtca tgagctgggt ccgccaggct 120ccagggaagg ggctggagtg ggtctcagct attagtgcta gtggtggtag tatatactac 180gcagactccg tgaagggccg gttcaccatc tccagagaca attccaagaa cacgctgtat 240ctgcaaatga acagcctgag agccgaggac acggccgtat attactgtgc gaaatccccg 300tataactgga acccctttga ctattggggc cagggaaccc tggtcaccgt ctcctca 357142119PRTArtificial SequenceSynthetic 142Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Leu Asn Asn Phe 20 25 30 Val Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Ala Ser Gly Gly Ser Ile Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Ser Pro Tyr Asn Trp Asn Pro Phe Asp Tyr Trp Gly Gln Gly 100 105 110 Thr Leu Val Thr Val Ser Ser 115 143325DNAArtificial SequenceSynthetic 143gaaatagtga tgacgcagtc tccagccacc ctgtctgtgt ctccagggga aagagccacc 60ctctcctgca gggccagtct gagtgttagc agcaaattag cctggtacca gcagaaacct 120ggccaggctc ccaggctcct catctatagt gcctccacca gggccactgg tatcccagcc 180aggttcagtg gcagtgggtc tgggacagag ttcactctca ccatcagcag cctgcagtct 240gaagattttg cagtttatta ctgtcagcag tataatcatt ggcctccgta cacttttggc 300caggggacca agctggagat caaac 325144108PRTArtificial SequenceSynthetic 144Glu Ile Val Met Thr Gln Ser Pro Ala Thr Leu Ser Val Ser Pro Gly1 5 10 15 Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Leu Ser Val Ser Ser Lys 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile 35 40 45 Tyr Ser Ala Ser Thr Arg Ala Thr Gly Ile Pro Ala Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Glu Phe Thr Leu Thr Ile Ser Ser Leu Gln Ser65 70 75 80 Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Tyr Asn His Trp Pro Pro 85 90 95 Tyr Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 145375DNAArtificial SequenceSynthetic 145caggtgcagc tggtggagtc tgggggaggc ttggaacagc cgggggggtc cctgagactc 60tcctgtgcag gctctggatt cacctttaga gactatgcca tgacctgggt ccgccaggct 120ccagggaagg gactggagtg ggtctcatct attagtggtt ccggtggtaa cacatactac 180gcagactccg tgaagggccg gttcaccatc tccagagaca attccaagaa cacgctgtat 240ctgcaaatga acagcctgag agccgaggac acggccgtat attactgtgc gaaagatcga 300ctctctataa caattcgccc acgctattat ggtttggacg tctggggcca agggtccacg 360gtcaccgtct cctca 375146125PRTArtificial SequenceSynthetic 146Gln Val Gln Leu Val Glu Ser Gly Gly Gly Leu Glu Gln Pro Gly Gly1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Gly Ser Gly Phe Thr Phe Arg Asp Tyr 20 25 30 Ala Met Thr Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Ile Ser Gly Ser Gly Gly Asn Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Asp Arg Leu Ser Ile Thr Ile Arg Pro Arg Tyr Tyr Gly Leu 100 105 110 Asp Val Trp Gly Gln Gly Ser Thr Val Thr Val Ser Ser 115 120 125 14724DNAArtificial SequenceSynthetic 147ggattcacct ttagagacta tgcc 241488PRTArtificial SequenceSynthetic 148Gly Phe Thr Phe Arg Asp Tyr Ala1 5 14924DNAArtificial SequenceSynthetic 149attagtggtt ccggtggtaa caca 241508PRTArtificial SequenceSynthetic 150Ile Ser Gly Ser Gly Gly Asn Thr1 5 15154DNAArtificial SequenceSynthetic 151gcgaaagatc gactctctat aacaattcgc ccacgctatt atggtttgga cgtc 5415218PRTArtificial SequenceSynthetic 152Ala Lys Asp Arg Leu Ser Ile Thr Ile Arg Pro Arg Tyr Tyr Gly Leu1 5 10 15 Asp Val153339DNAArtificial SequenceSynthetic 153gacatcgtgt tgacccagtc tccactctcc ctgcccgtca cccctggaga gccggcctcc 60atctcctgca ggtctagtca gagcctcctg tatagtattg gatacaacta tttggattgg 120tacctgcaga agtcagggca gtctccacag ctccttatct atttgggttc taatcgggcc 180tccggggtcc ctgacaggtt cagtggcagt ggatcaggca cagattttac actgaaaatc 240agcagagtgg aggctgagga tgttgggttt tattactgca tgcaagctct acaaactccg 300tacacttttg gccaggggac caagctggag atcaaacga 339154113PRTArtificial SequenceSynthetic 154Asp Ile Val Leu Thr Gln Ser Pro Leu Ser Leu Pro Val Thr Pro Gly1 5 10 15 Glu Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Leu Leu Tyr Ser 20 25 30 Ile Gly Tyr Asn Tyr Leu Asp Trp Tyr Leu Gln Lys Ser Gly Gln Ser 35 40 45 Pro Gln Leu Leu Ile Tyr Leu Gly Ser Asn Arg Ala Ser Gly Val Pro 50 55 60 Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile65 70 75 80 Ser Arg Val Glu Ala Glu Asp Val Gly Phe Tyr Tyr Cys Met Gln Ala 85 90 95 Leu Gln Thr Pro Tyr Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 110 Arg 15533DNAArtificial SequenceSynthetic 155cagagcctcc tgtatagtat tggatacaac tat 3315611PRTArtificial SequenceSynthetic 156Gln Ser Leu Leu Tyr Ser Ile Gly Tyr Asn Tyr1 5 10 1579DNAArtificial SequenceSynthetic 157ttgggttct 91583PRTArtificial SequenceSynthetic 158Leu Gly Ser1 15927DNAArtificial SequenceSynthetic 159atgcaagctc tacaaactcc gtacact 271609PRTArtificial SequenceSynthetic 160Met Gln Ala Leu Gln Thr Pro Tyr Thr1 5 161372DNAArtificial SequenceSynthetic 161gaggtgcagc tggtggagtc tgggggaggc ttggaacagc cgggggggtc cctgagactc 60tcctgtgcag gctctggatt cacctttaga gactatgcca tgacctgggt ccgccaggct 120ccagggaagg gactggagtg ggtctcatct attagtggtt ccggtggtaa cacatactac 180gcagactccg tgaagggccg gttcaccatc tccagagaca attccaagaa cacgctgtat 240ctgcaaatga acagcctgag agccgaggac acggccgtat attactgtgc gaaagatcga 300ctctctataa caattcgccc acgctattat ggtttggacg tctggggcca agggaccacg 360gtcaccgtct cc 372162124PRTArtificial SequenceSynthetic 162Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Glu Gln Pro Gly Gly1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Gly Ser Gly Phe Thr Phe Arg Asp Tyr 20 25 30 Ala Met Thr Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Ile Ser Gly Ser Gly Gly Asn Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Asp Arg Leu Ser Ile Thr Ile Arg Pro Arg Tyr Tyr Gly Leu 100 105 110 Asp Val Trp Gly Gln Gly Thr Thr Val Thr Val Ser 115 120 163336DNAArtificial SequenceSynthetic 163gacatcgtga tgacccagtc tccactctcc ctgcccgtca cccctggaga gccggcctcc 60atctcctgca ggtctagtca gagcctcctg tatagtattg gatacaacta tttggattgg 120tacctgcaga agtcagggca gtctccacag ctccttatct atttgggttc taatcgggcc 180tccggggtcc ctgacaggtt cagtggcagt ggatcaggca cagattttac actgaaaatc 240agcagagtgg aggctgagga tgttgggttt tattactgca tgcaagctct acaaactccg 300tacacttttg gccaggggac caagctggag atcaaa 336164112PRTArtificial SequenceSynthetic 164Asp Ile Val Met Thr Gln Ser Pro Leu Ser Leu Pro Val Thr Pro Gly1 5 10 15 Glu Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Leu Leu Tyr Ser 20 25 30 Ile Gly Tyr Asn Tyr Leu Asp Trp Tyr Leu Gln Lys Ser Gly Gln Ser 35 40 45 Pro Gln Leu Leu Ile Tyr Leu Gly Ser Asn Arg Ala Ser Gly Val Pro 50 55 60 Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile65 70 75 80 Ser Arg Val Glu Ala Glu Asp Val Gly Phe Tyr Tyr Cys Met Gln Ala 85 90 95 Leu Gln Thr Pro Tyr Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 110 165373DNAArtificial SequenceSynthetic 165gaggtgcagc tggtggagtc tgggggaggc ttggtacagc ctggggggtc cctgagactc 60tcctgtgcag cctctggatt cacctttaga gactatgcca tgagctgggt ccgccaggct 120ccagggaagg ggctggagtg ggtctcagct attagtggtt ccggtggtaa cacatactac 180gcagactccg tgaagggccg gttcaccatc tccagagaca attccaagaa cacgctgtat 240ctgcaaatga acagcctgag agccgaggac acggccgtat attactgtgc gaaagatcga 300ctctctataa caattcgccc acgctattat ggtttggacg tctggggcca agggaccacg 360gtcaccgtct cct 373166124PRTArtificial SequenceSynthetic 166Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Arg Asp Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Gly Ser Gly Gly Asn Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Asp Arg Leu Ser Ile Thr Ile Arg Pro Arg Tyr Tyr Gly Leu 100 105 110 Asp Val Trp Gly Gln Gly Thr Thr Val Thr Val Ser 115 120 167337DNAArtificial SequenceSynthetic 167gatattgtga tgactcagtc tccactctcc ctgcccgtca cccctggaga gccggcctcc 60atctcctgca ggtctagtca gagcctcctg tatagtattg gatacaacta tttggattgg 120tacctgcaga agccagggca gtctccacag ctcctgatct atttgggttc taatcgggcc 180tccggggtcc ctgacaggtt cagtggcagt ggatcaggca cagattttac actgaaaatc 240agcagagtgg aggctgagga tgttggggtt tattactgca tgcaagctct acaaactccg 300tacacttttg gccaggggac caagctggag atcaaac 337168112PRTArtificial SequenceSynthetic 168Asp Ile Val Met Thr Gln Ser Pro Leu Ser Leu Pro Val Thr Pro Gly1 5 10 15 Glu Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Leu Leu Tyr Ser 20 25 30 Ile Gly Tyr Asn Tyr Leu Asp Trp Tyr Leu Gln Lys Pro Gly Gln Ser 35 40 45 Pro Gln Leu Leu Ile Tyr Leu Gly Ser Asn Arg Ala Ser Gly Val Pro 50 55 60 Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile65 70 75 80 Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Met Gln Ala 85 90 95 Leu Gln Thr Pro Tyr Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 110 169375DNAArtificial SequenceSynthetic 169caggtgcagc tggtggagtc tgggggagtc ttggagcagc ctggggggtc cctgagactc 60tcctgtacag cctctggatt cacctttaga gactatgcca tgacctgggt ccgccaggct 120ccagggaagg ggctggagtg ggtctcatct attagtggta gtggtggtaa tacatactac 180gcagactccg tgaggggccg gttcaccatc tccagagaca actccaacca cacgctgtat 240ctgcaaatga acagcctgag agccgaagac acggccgtat attactgtgc gaaagatcga 300ctctccataa caattcgccc acgctattac ggtttggacg tctggggcca agggtccacg 360gtcaccgtct cctca 375170125PRTArtificial SequenceSynthetic 170Gln Val Gln Leu Val Glu Ser Gly Gly Val Leu Glu Gln Pro Gly Gly1 5 10 15 Ser Leu Arg Leu Ser Cys Thr Ala Ser Gly Phe Thr Phe Arg Asp Tyr 20 25 30 Ala Met Thr Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Ile Ser Gly Ser Gly Gly Asn Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Arg Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Asn His Thr Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Asp Arg Leu Ser Ile Thr Ile Arg Pro Arg Tyr Tyr Gly Leu 100 105 110 Asp Val Trp Gly Gln Gly Ser Thr Val Thr Val Ser Ser 115 120 125 17124DNAArtificial SequenceSynthetic 171ggattcacct ttagagacta tgcc 241728PRTArtificial SequenceSynthetic 172Gly Phe Thr Phe Arg Asp Tyr Ala1 5 17324DNAArtificial SequenceSynthetic 173attagtggta gtggtggtaa taca 241748PRTArtificial SequenceSynthetic 174Ile Ser Gly Ser Gly Gly Asn Thr1 5 17554DNAArtificial SequenceSynthetic 175gcgaaagatc gactctccat aacaattcgc ccacgctatt acggtttgga cgtc 5417618PRTArtificial SequenceSynthetic 176Ala Lys Asp Arg Leu Ser Ile Thr Ile Arg Pro Arg Tyr Tyr Gly Leu1 5 10 15 Asp Val177324DNAArtificial SequenceSynthetic 177gatattgtga tgacccagtc tccatcctcc ctgtctgcat ctgtaggaga cagagtcacc 60attacttgcc gggcgagtca ggacattagc aattattttg cctggtatca gcagaagcca 120gggaaagttc ctaaactcct gatctttgct gcatccactt tgcatccagg ggtcccatct 180cggttcagtg gcagtggatc tgggacagat ttcactctca ccattagtag cctgcagcct 240gaagatgttg caacttatta ctgtcaaaag tataacagtg ccccgtacac ttttggccag 300gggaccaagg tggaaatcaa

acga 324178108PRTArtificial SequenceSynthetic 178Asp Ile Val Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Ile Ser Asn Tyr 20 25 30 Phe Ala Trp Tyr Gln Gln Lys Pro Gly Lys Val Pro Lys Leu Leu Ile 35 40 45 Phe Ala Ala Ser Thr Leu His Pro Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro65 70 75 80 Glu Asp Val Ala Thr Tyr Tyr Cys Gln Lys Tyr Asn Ser Ala Pro Tyr 85 90 95 Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Arg 100 105 17918DNAArtificial SequenceSynthetic 179caggacatta gcaattat 181806PRTArtificial SequenceSynthetic 180Gln Asp Ile Ser Asn Tyr1 5 1819DNAArtificial SequenceSynthetic 181gctgcatcc 91823PRTArtificial SequenceSynthetic 182Ala Ala Ser1 18327DNAArtificial SequenceSynthetic 183caaaagtata acagtgcccc gtacact 271849PRTArtificial SequenceSynthetic 184Gln Lys Tyr Asn Ser Ala Pro Tyr Thr1 5 185372DNAArtificial SequenceSynthetic 185gaggtgcagc tggtggagtc tgggggagtc ttggagcagc ctggggggtc cctgagactc 60tcctgtacag cctctggatt cacctttaga gactatgcca tgacctgggt ccgccaggct 120ccagggaagg ggctggagtg ggtctcatct attagtggta gtggtggtaa tacatactac 180gcagactccg tgaggggccg gttcaccatc tccagagaca actccaacca cacgctgtat 240ctgcaaatga acagcctgag agccgaagac acggccgtat attactgtgc gaaagatcga 300ctctccataa caattcgccc acgctattac ggtttggacg tctggggcca agggaccacg 360gtcaccgtct cc 372186124PRTArtificial SequenceSynthetic 186Glu Val Gln Leu Val Glu Ser Gly Gly Val Leu Glu Gln Pro Gly Gly1 5 10 15 Ser Leu Arg Leu Ser Cys Thr Ala Ser Gly Phe Thr Phe Arg Asp Tyr 20 25 30 Ala Met Thr Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Ile Ser Gly Ser Gly Gly Asn Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Arg Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Asn His Thr Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Asp Arg Leu Ser Ile Thr Ile Arg Pro Arg Tyr Tyr Gly Leu 100 105 110 Asp Val Trp Gly Gln Gly Thr Thr Val Thr Val Ser 115 120 187321DNAArtificial SequenceSynthetic 187gacatccaga tgacccagtc tccatcctcc ctgtctgcat ctgtaggaga cagagtcacc 60attacttgcc gggcgagtca ggacattagc aattattttg cctggtatca gcagaagcca 120gggaaagttc ctaaactcct gatctttgct gcatccactt tgcatccagg ggtcccatct 180cggttcagtg gcagtggatc tgggacagat ttcactctca ccattagtag cctgcagcct 240gaagatgttg caacttatta ctgtcaaaag tataacagtg ccccgtacac ttttggccag 300gggaccaagc tggagatcaa a 321188107PRTArtificial SequenceSynthetic 188Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Ile Ser Asn Tyr 20 25 30 Phe Ala Trp Tyr Gln Gln Lys Pro Gly Lys Val Pro Lys Leu Leu Ile 35 40 45 Phe Ala Ala Ser Thr Leu His Pro Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro65 70 75 80 Glu Asp Val Ala Thr Tyr Tyr Cys Gln Lys Tyr Asn Ser Ala Pro Tyr 85 90 95 Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 189373DNAArtificial SequenceSynthetic 189gaggtgcagc tggtggagtc tgggggaggc ttggtacagc ctggggggtc cctgagactc 60tcctgtgcag cctctggatt cacctttaga gactatgcca tgagctgggt ccgccaggct 120ccagggaagg ggctggagtg ggtctcagct attagtggta gtggtggtaa tacatactac 180gcagactccg tgaagggccg gttcaccatc tccagagaca attccaagaa cacgctgtat 240ctgcaaatga acagcctgag agccgaggac acggccgtat attactgtgc gaaagatcga 300ctctccataa caattcgccc acgctattac ggtttggacg tctggggcca agggaccacg 360gtcaccgtct cct 373190124PRTArtificial SequenceSynthetic 190Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Arg Asp Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Gly Ser Gly Gly Asn Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Asp Arg Leu Ser Ile Thr Ile Arg Pro Arg Tyr Tyr Gly Leu 100 105 110 Asp Val Trp Gly Gln Gly Thr Thr Val Thr Val Ser 115 120 191322DNAArtificial SequenceSynthetic 191gacatccaga tgacccagtc tccatcctcc ctgtctgcat ctgtaggaga cagagtcacc 60atcacttgcc gggcgagtca ggacattagc aattatttag cctggtatca gcagaaacca 120gggaaagttc ctaagctcct gatctatgct gcatccactt tgcaatcagg ggtcccatct 180cggttcagtg gcagtggatc tgggacagat ttcactctca ccatcagcag cctgcagcct 240gaagatgttg caacttatta ctgtcaaaag tataacagtg ccccgtacac ttttggccag 300gggaccaagc tggagatcaa ac 322192107PRTArtificial SequenceSynthetic 192Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Ile Ser Asn Tyr 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Lys Val Pro Lys Leu Leu Ile 35 40 45 Tyr Ala Ala Ser Thr Leu Gln Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro65 70 75 80 Glu Asp Val Ala Thr Tyr Tyr Cys Gln Lys Tyr Asn Ser Ala Pro Tyr 85 90 95 Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 193355DNAArtificial SequenceSynthetic 193gaagtgcacc tggtggaatc tgggggaggc ttggtacagc ctggcaggtc cctgagactc 60tcctgtgagg cctctggatt cacctttgat gattatgcca tgcactgggt ccggcaagct 120ccggggaagg gcctggaatg ggtctcaggt cttagtcgga caagtgtcag tataggctat 180gcggactctg tgaagggccg attcaccatc tccagagaca acgccaagaa ctccctttat 240ttggaaatga acagtctgag acctgaggac acggccttat attactgtgc aaaatggggg 300acccgggggt attttgacta ctggggccag ggaaccctgg tcaccgtctc ctcag 355194118PRTArtificial SequenceSynthetic 194Glu Val His Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Arg1 5 10 15 Ser Leu Arg Leu Ser Cys Glu Ala Ser Gly Phe Thr Phe Asp Asp Tyr 20 25 30 Ala Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Gly Leu Ser Arg Thr Ser Val Ser Ile Gly Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Ser Leu Tyr65 70 75 80 Leu Glu Met Asn Ser Leu Arg Pro Glu Asp Thr Ala Leu Tyr Tyr Cys 85 90 95 Ala Lys Trp Gly Thr Arg Gly Tyr Phe Asp Tyr Trp Gly Gln Gly Thr 100 105 110 Leu Val Thr Val Ser Ser 115 19524DNAArtificial SequenceSynthetic 195ggattcacct ttgatgatta tgcc 241968PRTArtificial SequenceSynthetic 196Gly Phe Thr Phe Asp Asp Tyr Ala1 5 19724DNAArtificial SequenceSynthetic 197cttagtcgga caagtgtcag tata 241988PRTArtificial SequenceSynthetic 198Leu Ser Arg Thr Ser Val Ser Ile1 5 19933DNAArtificial SequenceSynthetic 199gcaaaatggg ggacccgggg gtattttgac tac 3320011PRTArtificial SequenceSynthetic 200Ala Lys Trp Gly Thr Arg Gly Tyr Phe Asp Tyr1 5 10 201322DNAArtificial SequenceSynthetic 201gacatccaga tgacccagtc tccatcttcc gtgtctgcat ctgtgggaga cagagtcacc 60atcacttgtc gggcgagtca ggatattagt atttggttag cctggtatca gcagagtcca 120gggaaagccc ctaaactcct gatcaatgtt gcatcccgtt tgcaaagtgg ggtcccatca 180aggttcagcg gcagtggatc tgggacagat ttcactctca ccatcaacag tctgcagcct 240gaagattttg taacttacta ttgtcaacag gctaacagtt tcccgatcac cttcggccaa 300gggacacgac tggcgaccaa ac 322202107PRTArtificial SequenceSynthetic 202Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Val Ser Ala Ser Val Gly1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Ile Ser Ile Trp 20 25 30 Leu Ala Trp Tyr Gln Gln Ser Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Asn Val Ala Ser Arg Leu Gln Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Asn Ser Leu Gln Pro65 70 75 80 Glu Asp Phe Val Thr Tyr Tyr Cys Gln Gln Ala Asn Ser Phe Pro Ile 85 90 95 Thr Phe Gly Gln Gly Thr Arg Leu Ala Thr Lys 100 105 20318DNAArtificial SequenceSynthetic 203caggatatta gtatttgg 182046PRTArtificial SequenceSynthetic 204Gln Asp Ile Ser Ile Trp1 5 2059DNAArtificial SequenceSynthetic 205gttgcatcc 92063PRTArtificial SequenceSynthetic 206Val Ala Ser1 20727DNAArtificial SequenceSynthetic 207caacaggcta acagtttccc gatcacc 272089PRTArtificial SequenceSynthetic 208Gln Gln Ala Asn Ser Phe Pro Ile Thr1 5 209355DNAArtificial SequenceSynthetic 209gaggtgcagc tggtggagtc tgggggaggc ttggtacagc ctggcaggtc cctgagactc 60tcctgtgagg cctctggatt cacctttgat gattatgcca tgcactgggt ccggcaagct 120ccggggaagg gcctggaatg ggtctcaggt cttagtcgga caagtgtcag tataggctat 180gcggactctg tgaagggccg attcaccatc tccagagaca acgccaagaa ctccctttat 240ttggaaatga acagtctgag acctgaggac acggccttat attactgtgc aaaatggggg 300acccgggggt attttgacta ctggggccag ggaaccctgg tcaccgtctc ctcag 355210118PRTArtificial SequenceSynthetic 210Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Arg1 5 10 15 Ser Leu Arg Leu Ser Cys Glu Ala Ser Gly Phe Thr Phe Asp Asp Tyr 20 25 30 Ala Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Gly Leu Ser Arg Thr Ser Val Ser Ile Gly Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Ser Leu Tyr65 70 75 80 Leu Glu Met Asn Ser Leu Arg Pro Glu Asp Thr Ala Leu Tyr Tyr Cys 85 90 95 Ala Lys Trp Gly Thr Arg Gly Tyr Phe Asp Tyr Trp Gly Gln Gly Thr 100 105 110 Leu Val Thr Val Ser Ser 115 211322DNAArtificial SequenceSynthetic 211gacatccaga tgacccagtc tccatcttcc gtgtctgcat ctgtgggaga cagagtcacc 60atcacttgtc gggcgagtca ggatattagt atttggttag cctggtatca gcagagtcca 120gggaaagccc ctaaactcct gatcaatgtt gcatcccgtt tgcaaagtgg ggtcccatca 180aggttcagcg gcagtggatc tgggacagat ttcactctca ccatcaacag tctgcagcct 240gaagattttg taacttacta ttgtcaacag gctaacagtt tcccgatcac cttcggccaa 300gggacacgac tggagattaa ac 322212107PRTArtificial SequenceSynthetic 212Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Val Ser Ala Ser Val Gly1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Ile Ser Ile Trp 20 25 30 Leu Ala Trp Tyr Gln Gln Ser Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Asn Val Ala Ser Arg Leu Gln Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Asn Ser Leu Gln Pro65 70 75 80 Glu Asp Phe Val Thr Tyr Tyr Cys Gln Gln Ala Asn Ser Phe Pro Ile 85 90 95 Thr Phe Gly Gln Gly Thr Arg Leu Glu Ile Lys 100 105 213355DNAArtificial SequenceSynthetic 213gaagtgcagc tggtggagtc tgggggaggc ttggtacagc ctggcaggtc cctgagactc 60tcctgtgcag cctctggatt cacctttgat gattatgcca tgcactgggt ccggcaagct 120ccagggaagg gcctggagtg ggtctcaggt cttagtcgga caagtgtcag tataggctat 180gcggactctg tgaagggccg attcaccatc tccagagaca acgccaagaa ctccctgtat 240ctgcaaatga acagtctgag agctgaggac acggccttgt attactgtgc aaaatggggg 300acccgggggt attttgacta ctggggccaa ggaaccctgg tcaccgtctc ctcag 355214118PRTArtificial SequenceSynthetic 214Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Arg1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Asp Asp Tyr 20 25 30 Ala Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Gly Leu Ser Arg Thr Ser Val Ser Ile Gly Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Ser Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Leu Tyr Tyr Cys 85 90 95 Ala Lys Trp Gly Thr Arg Gly Tyr Phe Asp Tyr Trp Gly Gln Gly Thr 100 105 110 Leu Val Thr Val Ser Ser 115 215322DNAArtificial SequenceSynthetic 215gacatccaga tgacccagtc tccatcttcc gtgtctgcat ctgtaggaga cagagtcacc 60atcacttgtc gggcgagtca ggatattagt atttggttag cctggtatca gcagaaacca 120gggaaagccc ctaagctcct gatctatgtt gcatccagtt tgcaaagtgg ggtcccatca 180aggttcagcg gcagtggatc tgggacagat ttcactctca ccatcagcag cctgcagcct 240gaagattttg caacttacta ttgtcaacag gctaacagtt tcccgatcac cttcggccaa 300gggacacgac tggagattaa ac 322216107PRTArtificial SequenceSynthetic 216Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Val Ser Ala Ser Val Gly1 5 10 15 Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Gln Asp Ile Ser Ile Trp 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Tyr Val Ala Ser Ser Leu Gln Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Gln Gln Ala Asn Ser Phe Pro Ile 85 90 95 Thr Phe Gly Gln Gly Thr Arg Leu Glu Ile Lys 100 105 217363DNAArtificial SequenceSynthetic 217gaggtgcagc tgttggagtc tgggggaggc ttgctacagc cgggggggtc cctgagactc 60tcctgtgcag cctctggaat cacctttagc acctatgcca tgagctgggt ccgtcaggct 120ccagggaggg ggctggagtg ggtctcagct attagtggta gtggtgatag cacatcctac 180gcagactccg tgaagggccg gttcaccagc tccagagaca attccaagaa cacgctgtat 240ctgcaaatga acagcctgag agccgaggac acggccgtat attactgtgc gaaagtcata 300gcagctcgtc ctcactggaa cttcgatctc tggggccgtg gcaccctggt cactgtctcc 360tca 363218121PRTArtificial SequenceSynthetic 218Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Leu Gln Pro Gly Gly1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Ile Thr Phe Ser Thr Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Arg Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser

Gly Ser Gly Asp Ser Thr Ser Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ser Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Val Ile Ala Ala Arg Pro His Trp Asn Phe Asp Leu Trp Gly 100 105 110 Arg Gly Thr Leu Val Thr Val Ser Ser 115 120 21924DNAArtificial SequenceSynthetic 219ggaatcacct ttagcaccta tgcc 242208PRTArtificial SequenceSynthetic 220Gly Ile Thr Phe Ser Thr Tyr Ala1 5 22124DNAArtificial SequenceSynthetic 221attagtggta gtggtgatag caca 242228PRTArtificial SequenceSynthetic 222Ile Ser Gly Ser Gly Asp Ser Thr1 5 22342DNAArtificial SequenceSynthetic 223gcgaaagtca tagcagctcg tcctcactgg aacttcgatc tc 4222414PRTArtificial SequenceSynthetic 224Ala Lys Val Ile Ala Ala Arg Pro His Trp Asn Phe Asp Leu1 5 10 225324DNAArtificial SequenceSynthetic 225gaaattgtgt tgacacagtc tccagccacc ctgtctttgt ctccagggga aagagccacc 60ctctcctgca gggccagtca gagtgttagt agatatttag cctggtatca acagaaacct 120ggccaggctc ccaggctcct catctatgat gcatccaaca gggccactgg catcccagcc 180aggttcagtg gcagtgggtc tgggacagac ttcactctca ccatcagcag cctagagcct 240gaagattttg gagtttatta ctgtcagcag cgtagtgact ggccgctcac tttcggcgga 300gggaccaagg tggagatcaa acgg 324226107PRTArtificial SequenceSynthetic 226Glu Ile Val Leu Thr Gln Ser Pro Ala Thr Leu Ser Leu Ser Pro Gly1 5 10 15 Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Arg Tyr 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile 35 40 45 Tyr Asp Ala Ser Asn Arg Ala Thr Gly Ile Pro Ala Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Glu Pro65 70 75 80 Glu Asp Phe Gly Val Tyr Tyr Cys Gln Gln Arg Ser Asp Trp Pro Leu 85 90 95 Thr Phe Gly Gly Gly Thr Lys Val Glu Ile Lys 100 105 22718DNAArtificial SequenceSynthetic 227cagagtgtta gtagatat 182286PRTArtificial SequenceSynthetic 228Gln Ser Val Ser Arg Tyr1 5 2299DNAArtificial SequenceSynthetic 229gatgcatcc 92303PRTArtificial SequenceSynthetic 230Asp Ala Ser1 23127DNAArtificial SequenceSynthetic 231cagcagcgta gtgactggcc gctcact 272329PRTArtificial SequenceSynthetic 232Gln Gln Arg Ser Asp Trp Pro Leu Thr1 5 233363DNAArtificial SequenceSynthetic 233gaggtgcagc tgttggagtc tgggggaggc ttgctacagc cgggggggtc cctgagactc 60tcctgtgcag cctctggaat cacctttagc acctatgcca tgagctgggt ccgtcaggct 120ccagggaggg ggctggagtg ggtctcagct attagtggta gtggtgatag cacatcctac 180gcagactccg tgaagggccg gttcaccagc tccagagaca attccaagaa cacgctgtat 240ctgcaaatga acagcctgag agccgaggac acggccgtat attactgtgc gaaagtcata 300gcagctcgtc ctcactggaa cttcgatctc tggggccgtg gcaccctggt cactgtctcc 360tca 363234121PRTArtificial SequenceSynthetic 234Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Leu Gln Pro Gly Gly1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Ile Thr Phe Ser Thr Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Arg Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Gly Ser Gly Asp Ser Thr Ser Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ser Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Val Ile Ala Ala Arg Pro His Trp Asn Phe Asp Leu Trp Gly 100 105 110 Arg Gly Thr Leu Val Thr Val Ser Ser 115 120 235324DNAArtificial SequenceSynthetic 235gaaattgtgt tgacacagtc tccagccacc ctgtctttgt ctccagggga aagagccacc 60ctctcctgca gggccagtca gagtgttagt agatatttag cctggtatca acagaaacct 120ggccaggctc ccaggctcct catctatgat gcatccaaca gggccactgg catcccagcc 180aggttcagtg gcagtgggtc tgggacagac ttcactctca ccatcagcag cctagagcct 240gaagattttg gagtttatta ctgtcagcag cgtagtgact ggccgctcac tttcggcgga 300gggaccaagg tggagatcaa acgg 324236107PRTArtificial SequenceSynthetic 236Glu Ile Val Leu Thr Gln Ser Pro Ala Thr Leu Ser Leu Ser Pro Gly1 5 10 15 Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Arg Tyr 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile 35 40 45 Tyr Asp Ala Ser Asn Arg Ala Thr Gly Ile Pro Ala Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Glu Pro65 70 75 80 Glu Asp Phe Gly Val Tyr Tyr Cys Gln Gln Arg Ser Asp Trp Pro Leu 85 90 95 Thr Phe Gly Gly Gly Thr Lys Val Glu Ile Lys 100 105 237363DNAArtificial SequenceSynthetic 237gaggtgcagc tgttggagtc tgggggaggc ttggtacagc cgggggggtc cctgagactc 60tcctgtgcag cctctggaat cacctttagc acctatgcca tgagctgggt ccgtcaggct 120ccagggaagg ggctggagtg ggtctcagct attagtggta gtggtgatag cacatactac 180gcagactccg tgaagggccg gttcaccatc tccagagaca attccaagaa cacgctgtat 240ctgcaaatga acagcctgag agccgaggac acggccgtat attactgtgc gaaagtcata 300gcagctcgtc ctcactggaa cttcgatctc tggggccgtg gcaccctggt cactgtctcc 360tca 363238121PRTArtificial SequenceSynthetic 238Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Ile Thr Phe Ser Thr Tyr 20 25 30 Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Gly Ser Gly Asp Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Val Ile Ala Ala Arg Pro His Trp Asn Phe Asp Leu Trp Gly 100 105 110 Arg Gly Thr Leu Val Thr Val Ser Ser 115 120 239324DNAArtificial SequenceSynthetic 239gaaattgtgt tgacacagtc tccagccacc ctgtctttgt ctccagggga aagagccacc 60ctctcctgca gggccagtca gagtgttagt agatatttag cctggtatca acagaaacct 120ggccaggctc ccaggctcct catctatgat gcatccaaca gggccactgg catcccagcc 180aggttcagtg gcagtgggtc tgggacagac ttcactctca ccatcagcag cctagagcct 240gaagattttg cagtttatta ctgtcagcag cgtagtgact ggccgctcac tttcggcgga 300gggaccaagg tggagatcaa acgg 324240108PRTArtificial SequenceSynthetic 240Glu Ile Val Leu Thr Gln Ser Pro Ala Thr Leu Ser Leu Ser Pro Gly1 5 10 15 Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Arg Tyr 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile 35 40 45 Tyr Asp Ala Ser Asn Arg Ala Thr Gly Ile Pro Ala Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Glu Pro65 70 75 80 Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Arg Ser Asp Trp Pro Leu 85 90 95 Thr Phe Gly Gly Gly Thr Lys Val Glu Ile Lys Arg 100 105 241366DNAArtificial SequenceSynthetic 241caggtgcagc tggtggagtc tgggggaggc gtggtccagc ctgggaggtc cctgagactc 60acctgtgcag cctctggatt caccttcagt agtaatggca tgcactgggt ccgccaggct 120ccaggcaagg ggctggagtg ggtggcaatt atatcatatg atggaaataa tcaatactat 180gcagactccg tgaagggccg attcaccatc tccagagaca attccaagca cacgctgtat 240ctggaaatga acagcctgag agctgaggac acggctgtgt attactgtac aaaagccatc 300tctataagtg gaacttacaa ctggttcgat tcctggggcc agggaaccct ggtcaccgtc 360tcctca 366242122PRTArtificial SequenceSynthetic 242Gln Val Gln Leu Val Glu Ser Gly Gly Gly Val Val Gln Pro Gly Arg1 5 10 15 Ser Leu Arg Leu Thr Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Asn 20 25 30 Gly Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Ile Ile Ser Tyr Asp Gly Asn Asn Gln Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys His Thr Leu Tyr65 70 75 80 Leu Glu Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Thr Lys Ala Ile Ser Ile Ser Gly Thr Tyr Asn Trp Phe Asp Ser Trp 100 105 110 Gly Gln Gly Thr Leu Val Thr Val Ser Ser 115 120 24324DNAArtificial SequenceSynthetic 243ggattcacct tcagtagtaa tggc 242448PRTArtificial SequenceSynthetic 244Gly Phe Thr Phe Ser Ser Asn Gly1 5 24524DNAArtificial SequenceSynthetic 245atatcatatg atggaaataa tcaa 242468PRTArtificial SequenceSynthetic 246Ile Ser Tyr Asp Gly Asn Asn Gln1 5 24745DNAArtificial SequenceSynthetic 247acaaaagcca tctctataag tggaacttac aactggttcg attcc 4524815PRTArtificial SequenceSynthetic 248Thr Lys Ala Ile Ser Ile Ser Gly Thr Tyr Asn Trp Phe Asp Ser1 5 10 15 249324DNAArtificial SequenceSynthetic 249gaaattgtat tgacacagtc tccagccatc ctgtctttgt ctccagggga aagagccacc 60ctctcctgca gggccagtca gagtgttagc aggtacttag cctggtacca acagaaacct 120ggccaggctc ccaggctcct catctatgat gcatccaaca gggccactgg catcccagcc 180aggttcagtg gcagtgggtc tgggacagac ttcactctca ccatcagcag cctagagcct 240gaagattttg cagtttatta ctgtcaacag cgtagcaact ggccgctcac tttcggcgga 300gggaccaagg tggagatcaa acgg 324250107PRTArtificial SequenceSynthetic 250Glu Ile Val Leu Thr Gln Ser Pro Ala Ile Leu Ser Leu Ser Pro Gly1 5 10 15 Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Arg Tyr 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile 35 40 45 Tyr Asp Ala Ser Asn Arg Ala Thr Gly Ile Pro Ala Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Glu Pro65 70 75 80 Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Arg Ser Asn Trp Pro Leu 85 90 95 Thr Phe Gly Gly Gly Thr Lys Val Glu Ile Lys 100 105 25118DNAArtificial SequenceSynthetic 251cagagtgtta gcaggtac 182526PRTArtificial SequenceSynthetic 252Gln Ser Val Ser Arg Tyr1 5 2539DNAArtificial SequenceSynthetic 253gatgcatcc 92543PRTArtificial SequenceSynthetic 254Asp Ala Ser1 25527DNAArtificial SequenceSynthetic 255caacagcgta gcaactggcc gctcact 272569PRTArtificial SequenceSynthetic 256Gln Gln Arg Ser Asn Trp Pro Leu Thr1 5 257366DNAArtificial SequenceSynthetic 257caggtgcagc tggtggagtc tgggggaggc gtggtccagc ctgggaggtc cctgagactc 60acctgtgcag cctctggatt caccttcagt agtaatggca tgcactgggt ccgccaggct 120ccaggcaagg ggctggagtg ggtggcaatt atatcatatg atggaaataa tcaatactat 180gcagactccg tgaagggccg attcaccatc tccagagaca attccaagca cacgctgtat 240ctggaaatga acagcctgag agctgaggac acggctgtgt attactgtac aaaagccatc 300tctataagtg gaacttacaa ctggttcgat tcctggggcc agggaaccct ggtcaccgtc 360tcctca 366258122PRTArtificial SequenceSynthetic 258Gln Val Gln Leu Val Glu Ser Gly Gly Gly Val Val Gln Pro Gly Arg1 5 10 15 Ser Leu Arg Leu Thr Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Asn 20 25 30 Gly Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Ile Ile Ser Tyr Asp Gly Asn Asn Gln Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys His Thr Leu Tyr65 70 75 80 Leu Glu Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Thr Lys Ala Ile Ser Ile Ser Gly Thr Tyr Asn Trp Phe Asp Ser Trp 100 105 110 Gly Gln Gly Thr Leu Val Thr Val Ser Ser 115 120 259324DNAArtificial SequenceSynthetic 259gaaattgtat tgacacagtc tccagccatc ctgtctttgt ctccagggga aagagccacc 60ctctcctgca gggccagtca gagtgttagc aggtacttag cctggtacca acagaaacct 120ggccaggctc ccaggctcct catctatgat gcatccaaca gggccactgg catcccagcc 180aggttcagtg gcagtgggtc tgggacagac ttcactctca ccatcagcag cctagagcct 240gaagattttg cagtttatta ctgtcaacag cgtagcaact ggccgctcac tttcggcgga 300gggaccaagg tggagatcaa acgg 324260107PRTArtificial SequenceSynthetic 260Glu Ile Val Leu Thr Gln Ser Pro Ala Ile Leu Ser Leu Ser Pro Gly1 5 10 15 Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Arg Tyr 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile 35 40 45 Tyr Asp Ala Ser Asn Arg Ala Thr Gly Ile Pro Ala Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Glu Pro65 70 75 80 Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Arg Ser Asn Trp Pro Leu 85 90 95 Thr Phe Gly Gly Gly Thr Lys Val Glu Ile Lys 100 105 261366DNAArtificial SequenceSynthetic 261caggtgcagc tggtggagtc tgggggaggc gtggtccagc ctgggaggtc cctgagactc 60tcctgtgcag cctctggatt caccttcagt agtaatggca tgcactgggt ccgccaggct 120ccaggcaagg ggctggagtg ggtggcagtt atatcatatg atggaaataa tcaatactat 180gcagactccg tgaagggccg attcaccatc tccagagaca attccaagaa cacgctgtat 240ctgcaaatga acagcctgag agctgaggac acggctgtgt attactgtac aaaagccatc 300tctataagtg gaacttacaa ctggttcgat tcctggggcc agggaaccct ggtcaccgtc 360tcctca 366262122PRTArtificial SequenceSynthetic 262Gln Val Gln Leu Val Glu Ser Gly Gly Gly Val Val Gln Pro Gly Arg1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Asn 20 25 30 Gly Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ala Val Ile Ser Tyr Asp Gly Asn Asn Gln Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Thr Lys Ala Ile Ser Ile Ser Gly Thr Tyr Asn Trp Phe Asp Ser Trp 100 105 110 Gly Gln Gly Thr Leu Val Thr Val Ser Ser 115 120 263324DNAArtificial SequenceSynthetic 263gaaattgtat tgacacagtc tccagccacc ctgtctttgt ctccagggga aagagccacc 60ctctcctgca gggccagtca gagtgttagc aggtacttag cctggtacca acagaaacct 120ggccaggctc ccaggctcct catctatgat gcatccaaca gggccactgg catcccagcc 180aggttcagtg gcagtgggtc tgggacagac ttcactctca

ccatcagcag cctagagcct 240gaagattttg cagtttatta ctgtcaacag cgtagcaact ggccgctcac tttcggcgga 300gggaccaagg tggagatcaa acgg 324264108PRTArtificial SequenceSynthetic 264Glu Ile Val Leu Thr Gln Ser Pro Ala Thr Leu Ser Leu Ser Pro Gly1 5 10 15 Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Arg Tyr 20 25 30 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile 35 40 45 Tyr Asp Ala Ser Asn Arg Ala Thr Gly Ile Pro Ala Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Glu Pro65 70 75 80 Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Arg Ser Asn Trp Pro Leu 85 90 95 Thr Phe Gly Gly Gly Thr Lys Val Glu Ile Lys Arg 100 105 2658PRTArtificial SequenceSyntheticVARIANT(1)...(8)Xaa = Any amino acid 265Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa1 5 2668PRTArtificial SequenceSyntheticVARIANT(1)...(8)Xaa = Any amino acid 266Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa1 5 26718PRTArtificial SequenceSyntheticVARIANT(1)...(18)Xaa = Any amino acid 267Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa1 5 10 15 Xaa Xaa26811PRTArtificial SequenceSyntheticVARIANT(1)...(11)Xaa = Any amino acid 268Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa1 5 10 2693PRTArtificial SequenceSyntheticVARIANT(1)...(3)Xaa = Any amino acid 269Xaa Xaa Xaa1 2709PRTArtificial SequenceSyntheticVARIANT(1)...(9)Xaa = Any amino acid 270Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa Xaa1 5 271330PRTArtificial SequenceSynthetic 271Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys1 5 10 15 Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30 Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45 Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser 50 55 60 Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr65 70 75 80 Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95 Lys Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys 100 105 110 Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro 115 120 125 Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys 130 135 140 Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp145 150 155 160 Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu 165 170 175 Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu 180 185 190 His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn 195 200 205 Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly 210 215 220 Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu225 230 235 240 Leu Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr 245 250 255 Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn 260 265 270 Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe 275 280 285 Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn 290 295 300 Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr305 310 315 320 Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys 325 330 272327PRTArtificial SequenceSynthetic 272Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Cys Ser Arg1 5 10 15 Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30 Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45 Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser 50 55 60 Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Lys Thr65 70 75 80 Tyr Thr Cys Asn Val Asp His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95 Arg Val Glu Ser Lys Tyr Gly Pro Pro Cys Pro Ser Cys Pro Ala Pro 100 105 110 Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys 115 120 125 Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val 130 135 140 Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp Tyr Val Asp145 150 155 160 Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Phe 165 170 175 Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp 180 185 190 Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Gly Leu 195 200 205 Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg 210 215 220 Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Gln Glu Glu Met Thr Lys225 230 235 240 Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp 245 250 255 Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys 260 265 270 Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser 275 280 285 Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn Val Phe Ser 290 295 300 Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser305 310 315 320 Leu Ser Leu Ser Leu Gly Lys 325 273327PRTArtificial SequenceSynthetic 273Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Cys Ser Arg1 5 10 15 Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30 Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45 Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser 50 55 60 Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Lys Thr65 70 75 80 Tyr Thr Cys Asn Val Asp His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95 Arg Val Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys Pro Ala Pro 100 105 110 Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys 115 120 125 Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val 130 135 140 Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp Tyr Val Asp145 150 155 160 Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Phe 165 170 175 Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp 180 185 190 Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Gly Leu 195 200 205 Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg 210 215 220 Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Gln Glu Glu Met Thr Lys225 230 235 240 Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp 245 250 255 Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys 260 265 270 Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser 275 280 285 Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn Val Phe Ser 290 295 300 Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser305 310 315 320 Leu Ser Leu Ser Leu Gly Lys 325 274207PRTHomo sapiens 274Met Lys Val Leu Gln Glu Pro Thr Cys Val Ser Asp Tyr Met Ser Ile1 5 10 15 Ser Thr Cys Glu Trp Lys Met Asn Gly Pro Thr Asn Cys Ser Thr Glu 20 25 30 Leu Arg Leu Leu Tyr Gln Leu Val Phe Leu Leu Ser Glu Ala His Thr 35 40 45 Cys Ile Pro Glu Asn Asn Gly Gly Ala Gly Cys Val Cys His Leu Leu 50 55 60 Met Asp Asp Val Val Ser Ala Asp Asn Tyr Thr Leu Asp Leu Trp Ala65 70 75 80 Gly Gln Gln Leu Leu Trp Lys Gly Ser Phe Lys Pro Ser Glu His Val 85 90 95 Lys Pro Arg Ala Pro Gly Asn Leu Thr Val His Thr Asn Val Ser Asp 100 105 110 Thr Leu Leu Leu Thr Trp Ser Asn Pro Tyr Pro Pro Asp Asn Tyr Leu 115 120 125 Tyr Asn His Leu Thr Tyr Ala Val Asn Ile Trp Ser Glu Asn Asp Pro 130 135 140 Ala Asp Phe Arg Ile Tyr Asn Val Thr Tyr Leu Glu Pro Ser Leu Arg145 150 155 160 Ile Ala Ala Ser Thr Leu Lys Ser Gly Ile Ser Tyr Arg Ala Arg Val 165 170 175 Arg Ala Trp Ala Gln Cys Tyr Asn Thr Thr Trp Ser Glu Trp Ser Pro 180 185 190 Ser Thr Lys Trp His Asn Ser Tyr Arg Glu Pro Phe Glu Gln His 195 200 205 275231PRTMacaca fasicularis 275Met Gly Trp Leu Cys Ser Gly Leu Leu Phe Pro Val Ser Cys Leu Val1 5 10 15 Leu Leu Gln Val Ala Ser Ser Gly Ser Met Lys Val Leu Gln Glu Pro 20 25 30 Thr Cys Val Ser Asp Tyr Met Ser Ile Ser Thr Cys Glu Trp Lys Met 35 40 45 Gly Gly Pro Thr Asn Cys Ser Ala Glu Leu Arg Leu Leu Tyr Gln Leu 50 55 60 Val Phe Gln Ser Ser Glu Thr His Thr Cys Val Pro Glu Asn Asn Gly65 70 75 80 Gly Val Gly Cys Val Cys His Leu Leu Met Asp Asp Val Val Ser Met 85 90 95 Asp Asn Tyr Thr Leu Asp Leu Trp Ala Gly Gln Gln Leu Leu Trp Lys 100 105 110 Gly Ser Phe Lys Pro Ser Glu His Val Lys Pro Arg Ala Pro Gly Asn 115 120 125 Leu Thr Val His Thr Asn Val Ser Asp Thr Val Leu Leu Thr Trp Ser 130 135 140 Asn Pro Tyr Pro Pro Asp Asn Tyr Leu Tyr Asn Asp Leu Thr Tyr Ala145 150 155 160 Val Asn Ile Trp Ser Glu Asn Asp Pro Ala Tyr Ser Arg Ile His Asn 165 170 175 Val Thr Tyr Leu Lys Pro Thr Leu Arg Ile Pro Ala Ser Thr Leu Lys 180 185 190 Ser Gly Ile Ser Tyr Arg Ala Arg Val Arg Ala Trp Ala Gln His Tyr 195 200 205 Asn Thr Thr Trp Ser Glu Trp Ser Pro Ser Thr Lys Trp Tyr Asn Ser 210 215 220 Tyr Arg Glu Pro Phe Glu Gln225 230

Claims

1. An antibody or antigen-binding fragment thereof that specifically binds human IL-4R, comprising three heavy chain complementarity determining region (HCDR) sequences comprising SEQ ID NO:148, 150, and 152, respectively, and three light chain complementarity determining region (LCDR) sequences comprising SEQ ID No:156, 158, and 160, respectively.

2. The antibody or antigen-binding fragment of claim 1, wherein the heavy and light chain CDR amino acid sequences (HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, LCDR3) are encoded by nucleotide sequences of SEQ ID NO:147, 149, 151, 155, 157, and 159, respectively.

3. The antibody or antigen-binding fragment of claim 1, wherein the antibody or antigen binding fragment comprises a heavy chain variable region (HCVR) having an amino acid sequence of SEQ ID NO:162 and a light chain variable region (LCVR) having an amino acid sequence of SEQ ID NO:164.

4. The antibody or antigen-binding fragment of claim 1, characterized by an affinity for hIL-4R (K.sub.D) of .ltoreq.300 pM.

5. The antibody or antigen-binding fragment of claim 1, characterized by an affinity for hIL-4R (K.sub.D) of .ltoreq.100 pM.

6. The antibody or antigen-binding fragment of claim 1, characterized by an affinity for hIL-4R (K.sub.D) of .ltoreq.50 pM.

7. The antibody or antigen-binding fragment of claim 1, wherein the antibody or fragment thereof cross-reacts with monkey IL-4R.

8. The antibody or antigen-binding fragment of claim 1 for use in the treatment of asthma or atopic dermatitis.

9. A nucleic acid molecule comprising a nucleic acid sequence encoding the antibody or antigen-binding fragment of claim 1.

10. A vector comprising the nucleic acid sequence of claim 9.

11. An isolated host cell comprising the vector of claim 10.

12. A therapeutic composition comprising the antibody or antigen-binding fragment of claim 1 and an acceptable carrier.

13. The therapeutic composition of claim 12, further comprising a second therapeutic agent selected from montelukast, pranlukast, zafirlukast, and rilonacept.

14. The antibody or antigen-binding fragment of claim 1 for use in the treatment of asthma.

15. The antibody or antigen-binding fragment of claim 1 for use in the treatment of atopic dermatitis.

16. The antibody or antigen-binding fragment of claim 1, wherein the antibody or antigen-binding fragment thereof is a human antibody, and wherein the antibody or antigen-binding fragment thereof is made in a transgenic mouse.

17. The antibody or antigen-binding fragment of claim 1, wherein the antibody or antigen-binding fragment is a human antibody, and wherein the antibody or antigen-binding fragment thereof is isolated from a hybridoma that is derived from a B cell that is isolated from a transgenic mouse.

18. A therapeutic composition comprising an antibody or antigen-binding fragment thereof that specifically binds human IL-4R, and a pharmaceutically acceptable carrier, wherein the antibody or antigen binding fragment comprises a heavy chain variable region (HCVR) having an amino acid sequence of SEQ ID NO:162 and a light chain variable region (LCVR) having an amino acid sequence of SEQ ID NO:164.

19. A therapeutic composition comprising an antibody or antigen-binding fragment thereof that specifically binds human IL-4R, and a pharmaceutically acceptable carrier, wherein the antibody or antigen binding fragment comprises heavy and light chain CDR amino acid sequences (HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, LCDR3) having the amino acid sequences of SEQ ID NO:148, 150, 152, 156, 158, and 160, respectively.

20. The therapeutic composition of claim 19 formulated for subcutaneous administration.