CA2502668C

CA2502668C - Fully human antibody fab fragments with human interferon-gamma neutralizing activity

Info

Publication number: CA2502668C
Application number: CA2502668A
Authority: CA
Inventors: Rajendra V. Deshpande; Mei-Mei Tsai
Original assignee: Amgen Inc
Current assignee: Amgen Inc
Priority date: 2002-10-17
Filing date: 2002-10-17
Publication date: 2013-07-30
Anticipated expiration: 2022-10-17
Also published as: AU2002347978A1; CA2502668A1; AU2002347978B2; EP1560595A4; EP1560595A2; MXPA05003949A; JP2006502739A

Abstract

Selective binding agents of interferon-gamma (IFN.UPSILON.) are provided by the invention. More particularly, the invention provides for antibodies and antigen binding domains which selectively bind to IFN.UPSILON. and may be used to prevent or treat conditions relating to autoimmune and inflammatory diseases such as rheumatoid arthritis, systemic lupus erythematosus and multiple sclerosis. Nucleic acid molecules encoding said antibodies and antigen binding domains, and expression vectors and host cells for the production of same are also provided.

Description

FULLY HUMAN ANTIBODY Fab FRAGMENTS WITH HUMAN
INTERFERON-GAMMA NEUTRALIZING ACTIVITY
Field of the Invention The invention relates to novel fully human antibody Fab fragments that bind to human interferon gamma (hIFNy), and inhibit its interaction with the cognate receptor, IFNy-R, and/or modify biological actions elicited by IFNy. More particularly, the invention relates to neutralizing Fab fragments (Fabs) isolated through hIFNy-affinity-selections of a phage displayed library containing unique Fab fragments, which were then converted into full-length human IgG
antibodies. These novel fully human antibodies to hIFNy, having the desired qualities of hIFNy-neutralizing activity, high affinity, and long half-life in vivo, may be used to prevent or treat various autoimmune and inflammatory diseases. Nucleic acid molecules, vectors and host cells for the production of the fully human Fabs of the invention are also provided.
Background of the Invention Antibodies have played an essential role in biopharmaceutical research and drug discovery efforts for many decades. The utility of antibodies as therapeutic agents for the treatment of human diseases has been idealized for many years due to their:
(a) long half-life in vivo; (b) ability to bind target(s) with high affinity and specificity; and (c) potential to mediate immune effector functions (such as complement fixation and antibody-dependent cellular cytotoxicity).
The reduction of the therapeutic antibody concept to practice was severely limited, however, until now, by the adverse immunogenicity of antibodies obtained from non-human species which restricted long-term clinical utility of the antibodies. Recent technological advances have provided new ways of overcoming these limitations by providing a means of obtaining fully human antibodies with less immunogenicity and a longer-term therapeutic potential.
Additionally, developments in combinatorial library methods and antibody-engineering have opened opportunities for modification of antibody-affinity, half-life, and/or effector functions.
One such technology, which employs filamentous phage-displayed, combinatorial libraries of antibody fragments fused to the phage coat protein (so-called "phage displayed library"), has been effectively used to discover antibodies with high affinity, specificity, and agonistic or antagonistic acitivity in vivo.
Human interferon gamma (hIFM) is a lymphokine produced by activated T-lymphocytes and natural killer cells. It manifests antiproliferative, antiviral and immunomodulatory activites and binds to hIFM-R, a heterodimeric receptor on most primary cells of the immune system; Langer et al., Immunology Today, 9:393 (1988), and triggers a cascade of events leading to inflammation. The antiviral and immunomodulatory activity of 'FM is known to have beneficial effects in a number of clinical conditions. However, there are many clinical settings in which IFN/-activity is known to have deleterious effects. For example, autoimmune dieseases are associated with high levels of hIFN/ in the blood, and there is now evidence suggesting that sequestration of IFN/ is associated with symptomatic relief of autoimmune diseases such as rheumatoid arthritis (RA), systemic lupus erythematosus (SLE) and multiple sclerosis (MS); see, e.g., Skurkovich et al., Intern. Journal of Immunotherapy, 14:23-32 (1998);
Gerez et al., Clin. Exp. Immunol., 109:296-303 (1997).
IFNy-activity has also been linked to such disease states as cachexia, septic shock and Crohn's disease.
Because blocking the interaction of hIFNy to its receptor represents the most upstream step of intervention in this regard, a fully human antibody with hIFNy-neutralizing activity represents an attractive therapeutic product candidate. It is an object of the present invention to employ the phage displayed library technology to identify antibodies using human interferon-gamma (hIFNy) as the therapeutic target.
Summary of the Invention The present invention provides for novel fully human antibody Fab fragments that bind to human interferon-gamma (hIFNy). In one embodiment, the fully human antibody Fab fragments bind to hIFNy in a manner that partially or completely inhibits the interaction of hIFNy with its cognate receptor, hIFNy-R, and thereby partially or completely inhibits hIFNy activity; that is, the antibody is an antagonist of hIFNy. Preferably, the hIFNy is mammalian hIFNy. More preferably, the hIFNy is human hIFNy which may be in soluble or cell surface associated forms, or fragments, derivatives and variants thereof.

3a Specific aspects of the invention include:
- an isolated human antibody that binds specifically to human interferon gamma, wherein the antibody comprises: (a) a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:34, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:44, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:54, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:1, a light chain CDR2 consisting of the amino acid sequence of SEQ ID NO:12, and a light chain CDR3 consisting of the amino acid sequence of SEQ
ID NO:23; (b) a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:35, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:45, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:55, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:2, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:13, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:24; (c) a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:35, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:45, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:56, a light chain CDR1 consisting of the amino acid sequence of SEQ ID
NO:3, a light chain CDR2 consisting of the amino acid sequence of SEQ ID NO:14, and a light chain CDR3 consisting of the amino 3b acid sequence of SEQ ID NO:25; (d) a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:36, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:46, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:57, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:4, a light chain CDR2 consisting of the amino acid sequence of SEQ ID NO:15, and a light chain CDR3 consisting of the amino acid sequence of SEQ
ID NO:26; (e) a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:37, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:47, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:58, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:5, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:16, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:27; (f) a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:38, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:48, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:59, a light chain CDR1 consisting of the amino acid sequence of SEQ ID
NO:6, a light chain CDR2 consisting of the amino acid sequence of SEQ ID NO:17, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:28; (g) a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:39, a heavy chain 3c complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:49, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:60, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:7, a light chain CDR2 consisting of the amino acid sequence of SEQ ID NO:18, and a light chain CDR3 consisting of the amino acid sequence of SEQ
ID NO:29; (h) a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:40, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:50, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:61, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:8, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:19, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:30; (i) a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:41, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:51, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:62, a light chain CDR1 consisting of the amino acid sequence of SEQ ID
NO:9, a light chain CDR2 consisting of the amino acid sequence of SEQ ID NO:20, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:31; (j) a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:42, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:52, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid 3d sequence of SEQ ID NO:63, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:10, a light chain CDR2 consisting of the amino acid sequence of SEQ ID NO:21, and a light chain CDR3 consisting of the amino acid sequence of SEQ
ID NO:32; or (k) a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID
NO:43, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:53, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:64, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:11, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:22, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:33;
a nucleic acid molecule encoding the antibody of the invention; and use of a therapeutically effective amount of the antibody of the invention for preventing or treating an autoimmune disease or an inflammatory condition.

An antibody of the present invention may be prepared by immunizing an animal with hIFNy such as murine or human hIFNy, preferably human hIFNy, or with an immunogenic fragment, derivative or variant thereof.
In addition, an animal may be immunized with cells transfected with a vector containing a nucleic acid molecule encoding hIFNy such that hIFNy is expressed and associated with the surface of the transfected cells. Alternatively, the antibodies may be obtained by screening a library comprising antibody or antigen binding domain sequences for binding to hIFNy. Such a library is conveniently prepared in bacteriophage as protein or peptide fusions to a bacteriophage coat protein which are expressed on the surface of assembled phage particles and the encoding DNA sequences contained within the phage particles (so-called "phage displayed library"). In one example, a phage displayed library contains DNA sequences encoding human antibodies, such as variable light and heavy chains.
The antibodies or antigen binding domains may be tetrameric glycoproteins similar to native antibodies, or they may be single chain antibodies; Fv, Fab, Fab' or F(ab)' fragments, bispecific antibodies, heteroantibodies, or other fragments, variants, or derivatives thereof, which are capable of binding hIFNy and partially or completely neutralize hIFNy activity.
Antibodies or antigen binding domains may be produced In hybridoma cell lines (antibody-producing cells such as spleen cells fused to mouse myeloma cells, for example) or may be produced in heterologous cell lines transfected with nucleic acid molecules encoding said antibody or antigen binding domain.
An antibody or antigen binding domain of the invention comprises:

(a) a Fab heavy chain amino acid sequence as shown in Figures 3-13 (SEQ ID NO:65-SEQ ID
NO:86);
(b) a heavy chain amino acid sequence comprising conservative amino acid substitutions of the sequence in (a);
(c) a heavy chain amino acid sequence which is at least about 80% identical to the sequence in (a); or (d) a fragment or derivative of (a), (b) or (c);
wherein the antibody or antigen binding domain binds selectively to hIFN.y.
In another embodiment, an antibody or antigen binding domain of the invention recognizes an epitope on human hIFNy recognized by an antibody or antigen binding domain comprising a Fab heavy chain amino acid sequence as shown in Figures 3-13 same as above (SEQ ID
NO:65-SEQ ID NO:86) and a Fab light amino acid sequence as shown in Figures 14-24 (SEQ ID NO:87-SEQ ID NO:108).
In another embodiment, an antibody or antigen binding domain of the invention comprises a V1 and Vh chain:
wherein each V, chain comprises CDR
amino acid sequences designated CDR1(V1), CDR2(V1) and CDR3(V1) separated by framework amino acid sequences, CDR1(V1) being selected from the group consisting of:
TGSSGSIASHYVQ (SEQ ID NO:01);
TGSSGSIASNYVQ (SEQ ID NO:02);
TRSSGSIASYYVQ (SEQ ID NO:03);
RATQSLLHGNGHNYLD (SEQ ID NO: 04);
RSSQSLVHSDGNTYLS (SEQ ID NO:05);
SGDVLARKYAR (SEQ ID NO:06);
GGDNLGGKSLH (SEQ ID NO: 07);
RSSQSLLHTNEYNYLD (SEQ ID NO:08);

TGSSGSIANNYVH (SEQ ID NO:09);
RASQYVSSNSLA (SEQ ID NO:10); and RSSQSLLRSNGYNYLA (SEQ ID NO:11) CDR2(V1) being selected from the group consisting of:
EDKERPS (SEQ ID NO:12);
EDNQRPS (SEQ ID NO:13);
EDDQRPS (SEQ ID NO:14);
MGSNRAS (SEQ ID NO:15);
KISNRFS (SEQ ID NO:16);
KDRERPS (SEQ ID NO:17);
DDSDRPS (SEQ ID NO:18);
LGSNRAP (SEQ ID NO:19);
EDDQRPS (SEQ ID NO:20);
GASNRAT (SEQ ID NO:21); and LASNRAS (SEQ ID NO:22) and CDR3(V1) being selected from the group consisting of:
QSYDSSNQWV (SEQ ID NO:23);
QSYDGSAWV (SEQ ID NO:24);
QSYDRNSLV (SEQ ID NO:25);
MQALQLPPT (SEQ ID NO:26);
MQATQLPYT (SEQ ID NO:27);
YSAADNRGV (SEQ ID NO: 28);
QVWDGSSDQRV (SEQ ID NO:29);
MQALQTPRT (SEQ ID NO: 30);
QSYDNSNSFVV (SEQ ID NO:31);
QQYGSSPIT (SEQ ID NO:32); AND
VHGVHIPYT (SEQ ID NO:33) wherein CDR1(V1), CDR2(V1) and CDR3(V1) are selected independently of each other; and wherein each Vh chain comprises CDR amino acid sequences designated CDR1(VJ, CDR2(VO and CDR3(VJ separated by framework amino acid sequences, CDR1(VJ being selected from the group consisting of:
GYYWS (SEQ ID NO:34);
SYAMS (SEQ ID NO:35);

GYYWS ( SEQ ID NO: 36 ) ;
NARMGVS ( SEQ ID NO : 37 ) ;
SYAMH ( SEQ ID NO: 38 ) ;
SYSMNT ( SEQ ID NO : 39) ;
GYYWS (SEQ ID NO:40);
SGGYSWS (SEQ ID NO:41);
SNYMS (SEQ ID NO:42); and SNEAGVG (SEQ ID NO:43) CDR2(Võ) being selected from the group consisting of:
EINHSGSTNYNPSLKS (SEQ ID NO:44);
AISGSGGSTYYADSVKG (SEQ ID NO:45);
EINHSGSTNYNPSLKS (SEQ ID NO:46);
HIFSNDEESYSTSLKS (SEQ ID NO:47);
VaSYDGSNKYYADSVKG (SEQ ID NO:48);
SISSGSSYRYDADSVKG (SEQ ID NO:49);
EINHSGSTNYNPSLKS (SEQ ID NO:50);
YIYHSGSTYYNPSLKS (SEQ ID NO:51);
VIYSGGSTYYADSVKG (SEQ ID NO:52); and LLYWDDDKRYSPSLRS (SEQ ID NO:53) CDR3(VJ being selected from the group consisting of:
GRARNWRSRFDY (SEQ ID NO: 54);
TSWNAGGPIDY (SEQ ID NO:55);
DRVGYSSSLLDY (SEQ ID NO:56);
DKGSRITIFGVVGSAGFDY (SEQ ID NO:57);
LLLYEGFDP (SEQ ID NO:58);
DLVLTMTSRRAAFDI (SEQ ID NO:59);
DQWGTISGNDY (SEQ ID NO:60);
GWPTYVWGSYRPKGYFDY (SEQ ID NO: 61);
GDWGYFDY (SEQ ID NO: 62);
DADGGDYGY (SEQ ID NO: 63); and RLVRYGGYSTGGFDV (SEQ ID NO:64) wherein CDR1(Võ), CDR2(VJ and CDR3(VJ are selected independently of each other.
In another embodiment, an antibody or antigen binding domain of the invention comprises a V1 and a V, chain wherein: the V, chain comprises CDR1 having the sequence TGSSGSIASHYVQ (SEQ ID NO:01) , CDR2 having the sequence EDKERPS (SEQ ID NO:12), and CDR3 having the sequence QSYDSSNQWV (SEQ ID NO:23); and the V, chain comprises CDR1 having the sequence GYYWS
(SEQ ID NO:34), CDR2 having the sequence EINHSGSTNYNPSLKS (SEQ ID NO:44), and CDR3 having the sequence GRARNWRSRFDY (SEQ ID NO:54);
wherein CDR1, CDR2 and CDR3 on each V, and V, chain are separated by framework amino acid sequences.
In another embodiment, an antibody or antigen binding domain of the invention comprises a V, and a V, chain wherein: the V, chain comprises CDR1 having the sequence TGSSGSIASNYVQ (SEQ ID NO:02), CDR2 having the sequence EDNQRPS (SEQ ID NO:13), and CDR3 having the sequence QSYDGSAWV (SEQ ID NO:24); and the V, chain comprises CDR1 having the sequence SYAMS
(SEQ ID NO:35), CDR2 having the sequence AISGSGGSTYYADSVKG (SEQ ID NO:45), and CDR3 having the sequence TSWNAGGPIDY (SEQ ID NO:55);
wherein CDR1, CDR2 and CDR3 on each V, and V, chain are separated by framework amino acid sequences.
In another embodiment, an antibody or antigen binding domain of the invention comprises a V, and a V, chain wherein: the V, chain comprises CDR1 having the sequence TRSSGSIASYYVQ (SEQ ID NO:03), CDR2 having the sequence EDDQRPS (SEQ ID NO:14), and CDR3 having the sequence QSYDRNSLV (SEQ ID NO:25); and the V, chain comprises CDR1 having the sequence SYAMS
(SEQ ID NO:35), CDR2 having the sequence AISGSGGSTYYADSVKG (SEQ ID NO: 45), and CDR3 having the sequence DRVGYSSSLLDY (SEQ ID NO:56);
wherein CDR1, CDR2 and CDR3 on each V, and V, chain are separated by framework amino acid sequences.
In another embodiment, an antibody or antigen binding domain of the invention comprises a V, and a V, chain wherein: the V, chain comprises CDR1 having the sequence RATQSLLHGNGHNYLD ( SEQ ID NO : 04 ) , CDR2 having the sequence MGSNRAS (SEQ ID NO: 15), and CDR3 having the sequence MQALQLPPT (SEQ ID NO:26); and the V, chain comprises CDR1 having the sequence GYYWS
(SEQ ID NO:36), CDR2 having the sequence EINHSGSTNYNPSLKS (SEQ ID NO: 46), and CDR3 having the sequence DKGSRITIFGVVGSAGFDY (SEQ ID NO: 57);
wherein CDR1, CDR2 and CDR3 on each V, and V, chain are separated by framework amino acid sequences.
In another embodiment, an antibody or antigen binding domain of the invention comprises a V, and a V, chain wherein: the V, chain comprises CDR1 having the sequence RSSQSLVHSDGNTYLS (SEQ ID NO:05), CDR2 having the sequence KISNRFS (SEQ ID NO: 16), and CDR3 having wherein CDR1, CDR2 and CDR3 on each V, and V, chain are separated by framework amino acid sequences.
In another embodiment, an antibody or antigen binding domain of the invention comprises a V, and a V, chain wherein: the V, chain comprises CDR1 having the VISYDGSNKYYADSVKG (SEQ ID NO:48), and CDR3 having the sequence DLVLTMTSRRAAFDI (SEQ ID NO: 59);
wherein CDR1, CDR2 and CDR3 on each V, and V, chain are separated by framework amino acid sequences.
In another embodiment, an antibody or antigen binding domain of the invention comprises a V, and a V, chain wherein: the V, chain comprises CDR1 having the sequence GGDNLGGKSLH (SEQ ID NO:0 7), CDR2 having the sequence DDSDRPS (SEQ ID NO:18), and CDR3 having the sequence QVWDGSSDQRV (SEQ ID NO:2 9); and the Vh chain comprises CDR1 having the sequence SYSMN
(SEQ ID NO:39), CDR2 having the sequence SISSGSSYRYDADSVKG (SEQ ID NO: 49), and CDR3 having the sequence DQWGTISGNDY (SEQ ID NO:60);
wherein CDR1, CDR2 and CDR3 on each V, and Vh chain are separated by framework amino acid sequences.
In another embodiment, an antibody or antigen binding domain of the invention comprises a V, and a Vh chain wherein: the V, chain comprises CDR1 having the sequence RSSQSLLHTNEYNYLD (SEQ ID NO:08), CDR2 having the sequence LGSNRAP (SEQ ID NO:19), and CDR3 having the sequence MQALQTPRT (SEQ ID NO: 30); and the Vh chain comprises CDR1 having the sequence GYYWS
(SEQ ID NO:40), CDR2 having the sequence EINHSGSTNYNPSLKS (SEQ ID NO:50), and CDR3 having the sequence GWPTYVWGSYRPKGYFDY (SEQ ID NO: 61);
wherein CDR1, CDR2 and CDR3 on each V, and Vh chain are separated by framework amino acid sequences.
In another embodiment, an antibody or antigen binding domain of the invention comprises a V, and a Vh chain wherein: the V, chain comprises CDR1 having the sequence TGSSGSIANNYVH (SEQ ID NO:09), CDR2 having the sequence EDDQRPS (SEQ ID NO:20), and CDR3 having the sequence QSYDNSNSFVV (SEQ ID NO:31); and the Vh chain comprises CDR1 having the sequence SGGYSWS
(SEQ ID NO:41), CDR2 having the sequence YIYHSGSTYYNPSLKS (SEQ ID NO:51), and CDR3 having the sequence GDWGYFDY (SEQ ID NO:62);
wherein CDR1, CDR2 and CDR3 on each V, and Vh chain are separated by framework amino acid sequences.
In another embodiment, an antibody or antigen binding domain of the invention comprises a V1 and a V, chain wherein: the V, chain comprises CDR1 having the sequence RASQYVSSNSLA (SEQ ID NO:10), CDR2 having the sequence GASNRAT (SEQ ID NO:21), and CDR3 having the sequence QQYGSSPIT (SEQ ID NO:32); and the Vh chain comprises CDR1 having the sequence SNYMS
(SEQ ID NO:42), CDR2 having the sequence VIYSGGSTYYADSVKG (SEQ ID NO: 52), and CDR3 having the sequence DADGGDYGY (SEQ ID NO: 63);
wherein CDR1, CDR2 and CDR3 on each V1 and Vh chain are separated by framework amino acid sequences.
In another embodiment, an antibody or antigen binding domain of the invention comprises a V, and a Vh chain wherein: the V1 chain comprises CDR1 having the sequence RSSQSLLRSNGYNYLA (SEQ ID NO:11), CDR2 having the sequence LASNRAS (SEQ ID NO:22), and CDR3 having the sequence VHGVHIPYT (SEQ ID NO:33); and the VI, chain comprises CDR1 having the sequence SNEAGVG
(SEQ ID NO:43), CDR2 having the sequence LLYWDDDKRYSPSLRS (SEQ ID NO:53), and CDR3 having the sequence RLVRYGGYSTGGFDV (SEQ ID NO: 64);
wherein CDR1, CDR2 and CDR3 on each V, and Vh chain are separated by framework amino acid sequences.
Antibodies and antigen binding domains of the invention are derived from germ line nucleic acid sequences present in genomic DNA which encode light and heavy chain amino acid sequences. Antibodies are encoded by nucleic acid sequences which are the products of germline sequence rearrangement and somatic mutation.
In one embodiment, an antibody or antigen binding domain of the invention comprises a V, and a VI, chain wherein the V, chain is comprises a rearranged or somatic variant of a VX6 germline genes such as in Figure 41 (SEQ ID NO:130); and the Vh chain comprises a rearranged or somatic variant of a VH4 germline genes such as in Figure 33 (SEQ ID NO:122); and the antibody binds selectively to an IFNy polypeptide.
In another embodiment, the V/ chain comprises or a rearranged or somatic variant of a VX6 germline genes such as in Figure 41 (SEQ ID NO:130); and the Vh chain comprises a rearranged or somatic variant of a VH1 germline gene such as in Figure 34 (SEQ ID NO:123).
In another embodiment, the V/ chain comprises a rearranged or somatic variant of a VK2 germline gene such as in Figure 42 (SEQ ID NO:131); and the Vh chain comprises a rearranged or somatic variant of a VH2 germline gene such as in Figure 35 (SEQ ID NO:124).
In another embodiment, the V/ chain comprises a rearranged or somatic variant of a VK2 germline gene such as in Figure 43 (SEQ ID NO:132); and the Vh chain comprises a rearranged or somatic variant of a VH4 germline gene such as in Figure 33 (SEQ ID NO:122).
In another embodiment, the V/ chain comprises a rearranged or somatic variant of a VX3 germline gene such as in Figure 44 (SEQ ID NO:133); and the Vh chain comprises a rearranged or somatic variant of a VH3 germline gene such as in Figure 36 (SEQ ID NO:125).
In another embodiment, the V1 chain comprises a rearranged or somatic variant of a VX3 germline gene such as in Figure 45 (SEQ ID NO:134); and the Vh chain comprises a rearranged or somatic variant of a VH3 germline gene such as in Figure 37 (SEQ ID NO:126).
In another embodiment, the V/ chain comprises a rearranged or somatic variant of a VK3 germline gene such as in Figure 46 (SEQ ID NO:135); and the Vh chain comprises a rearranged or somatic variant of a VH3 germline gene such as in Figure 38 (SEQ ID NO:127).
In another embodiment, the V/ chain comprises a rearranged or somatic variant of a VX6 germline gene such as in Figure 41 (SEQ ID NO:130); and the Vh chain comprises a rearranged or somatic variant of a VH4 germline gene such as in Figure 39 (SEQ ID NO:128).
In another embodiment, the V, chain comprises a rearranged or somatic variant of a Via germline gene such as in Figure 43 (SEQ ID NO:132); and the Vh chain comprises a rearranged or somatic variant of a VH4 germline gene such as in Figure 33 (SEQ ID NO:122).
In another embodiment, the V, chain comprises a rearranged or somatic variant of a V1(2 germline gene such as in Figure 43 (SEQ ID NO:132); and the Vh chain comprises a rearranged or somatic variant of a VH2 germline gene such as in Figure 40 (SEQ ID NO:129).
In another embodiment, the V1 chain comprises a rearranged or somatic variant of a VX6 germline gene such as in Figure 41 (SEQ ID NO:130); and the Vh chain comprises or a rearranged or somatic variant of a VH1 germline gene such as in Figure 34 (SEQ ID NO:123).
The selective binding agents of the invention (antibody or antigen binding domain) partially or completely inhibit at least one activity of 'FM, such as binding of IFNy to IFM-R.
In one embodiment, an IFN/ antagonist, such as an antibody or antigen binding domains, is administered to an animal which has experienced or is at risk of developing lupus-like disease, arthritis, or multiple-sclerosis-like syndrome. An IFNy antagonist may be used to prevent and/or treat lupus nephritis, rheumatoid arthritis, and/or multiple sclerosis.
Also provided are compositions comprising the antibodies or antigen binding domains of the invention and a pharmaceutically acceptable carrier.

Description of the Figures Figure 1 is a graph depicting the results of ELISA for reactivity of predominant phage Fab clones to hIFN.y. Phage dilutions were performed using a maximum of 100 41 of phage suspension pre-blocked with 2% MPBS
per well to given a typical range of 109-1011 phage/well in the ELISA. Phage stocks for ELISA were prepared as described in Example 3. Values were from single point determinations and ODa, was measured for signal detection.
Figure 2 is a graph depicting the results of a dose dependent clonal phage ELISA of predominant Fabs "GP-A" and "BS-B" clones for reactivity to hiFm.
Phage dilutions were performed using a maximum of 100 Al of phage suspension pre-blocked with 2% MPBS per well to given a typical range of 109-1011 phage/well in the ELISA. Phage stocks for ELISA were prepared as described in Example 3. Values were from single point determinations and OD,, was measured for signal detection.
Figure 3 shows the nucleotide and amino acid sequence of Fab "BS-A" heavy chain.
Figure 4 shows the nucleotide and amino acid sequence of Fab "BS-B" heavy chain.
Figure 5 shows the nucleotide and amino acid sequence of Fab "RD-B1" heavy chain.
Figure 6 shows the nucleotide and amino acid sequence of Fab "RD-A2" heavy chain.

Figure 7 shows the nucleotide and amino acid sequence of Fab "58C" heavy chain.
Figure 8 shows the nucleotide and amino acid sequence of Fab "GP-A" heavy chain.
Figure 9 shows the nucleotide and amino acid sequence of Fab "57D" heavy chain.
Figure 10 shows the nucleotide and amino acid sequence of Fab "57E" heavy chain.
Figure 11 shows the nucleotide and amino acid sequence of Fab "IFN-A" heavy chain.
Figure 12 shows the nucleotide and amino acid sequence of Fab "67C" heavy chain.
Figure 13 shows the nucleotide and amino acid sequence of Fab "59-A2" heavy chain.
Figure 14 shows the nucleotide and amino acid sequence of Fab "BS-A" light chain.
. Figure 15 shows the nucleotide and amino acid sequence of Fab "BS-B" light chain.
Figure 16 shows the nucleotide and amino acid sequence of Fab 1TRD-B1" light chain.
Figure 17 shows the nucleotide and amino acid sequence of Fab "RD-A2" light chain.
Figure 18 shows the nucleotide and amino acid sequence of Fab "58C" light chain.

Figure 19 shows the nucleotide and amino acid sequence of Fab "GP-A" light chain.
Figure 20 shows the nucleotide and amino acid sequence of Fab "57D" light chain.
Figure 21 shows the nucleotide and amino acid sequence of Fab "57E" light chain.
Figure 22 shows the nucleotide and amino acid sequence of Fab "IFN-A" light chain.
Figure 23 shows the nucleotide and amino acid sequence of Fab "67C" light chain.
Figure 24 shows the nucleotide and amino acid sequence of Fab "59-A2" light chain.
Figure 25 shows a comparison of the amino acid sequences of the heavy and light chain complementarily determining regions (CDRs) of Fabs "BS-A", "BS-B", "RD-A2",."RD-B1", "IFN-A", "57E", "57D", "GP-A", "58-C", "67C" and "59-A2".
Figure 26 is a graph depicting the neutralization activity of Fabs µ"BS-A" and "BS-B" as measured in the A549Tmce11 assay. Fabs were purified as described in Example 4 and added at Fab concentrations ranging from 0.3-150 Rg/ml. Pharmingen B27 Ab (concentrations ranging from 0.01-5 p.g/m1) was used as a positive control. Cells were stained with Alamar Blue 5 days post treatment, and analyzed 4 hours post staining on a FL50eplate reader.

Figure 27 is a graph depicting the neutralization activity of "BS-A" IgG and "BS-B" IgG as measured in the A549 cell assay. IgGs were purified as described in Example 4 and added at IgG concentrations ranging from 0.1-100 1.1,g/ml. Pharmingen B27 Ab (concentrations ranging from 0.01-5 glad) was used as a positive control. An irrelevant Ab, AT-IgG
(concentrations ranging from 0.01-5 ig/m1), that does not react with hIFNy was used as a negative control.
Cells were stained with Alamar Blue 5 days post treatment, and analyzed 4 hours post staining on a FL500 plate reader.
Figure 28 is a chart which provides a comparison of the affinity and neutralization activity of "BS-A", "BS-B", "RD-A2", "RD-B", "IFN-A", "57E", "57D", "GP-A", "58C" and "67C" IgGs as measured by BiaCore and in the A549 cell assay. The BiaCoremdata was analyzed using BIAEVALUATION.
Figure 29 is a graph depicting the neutralization activity of "BS-A" IgG and "BS-B" IgG as measured by BIACore. Relative binding response (%) is plotted vs. concentration of sample (nM).
Figure 30 is a chart which provides a comparison of affinity of anti-IFNy Fabs "BS-A", "BS-B", "IFN-A" and "GP-A" and the corresponding IgGs as measured by BIACore.
Figure 31 shows a comparison of Fab amino acid sequences shown in Figures 3-24. The predicted amino acid sequences of heavy and light chain Fabs "BS-A", "BS-B", "RD-A2", "RD-El", "IFN-A", "57E", "57D", "GP-A", "58-C", "67C" and "59-A2" were compared for identity and similarity. GCG's "BestFit" program was used to obtain percentage of identity and similarity between each pair of Fabs.
Figure 32 shows complementarily determining regions (CDRs) alignments of the heavy and light chain "BS-A", "BS-B", "RD-A2", "RD-B1", "IFN-A", "57E", "57D", "GP-A", "58-C", "67C" and "59-A2" Fabs.
Figure 33 shows a comparison of predicted Fab "BS-A", "RD-A2" and "IFN-A" heavy chain amino acid sequences (residues 1-120, 1-127 and 1-126 inclusive in Figures 3, 6 and 11, respectively) with germline sequence from the VH4 family. The germline sequence comprises the V region sequence 4-34, the D region sequences 1-1, 3-3 or 3-16, and the J region sequence JH4. FR1, FR2 and FR3 designate the three framework regions, CDR1, CDR2 and CDR3 designate the three complementarily determining regions, and H1, H2 and H3 designate the corresponding junction sequences between framework regions and CDRs. Differences between "BS-A", "RD-A2", "IFN-A" and germline V, D, or J sequences are in boldface. The numbering of germline amino acid residues in Figures 33-46 is as described in Kabat et al., Sequences of Proteins of Immunological Interest, U.S. Department of Health and Human Services, 4th ed.
(1991).
Figure 34 shows a comparison of predicted Fab "BS-B", and "59-A2" heavy chain amino acid sequences (residues 1-121 and 1-120 inclusive in Figures 4 and 13, respectively) with germline sequence from the VH1 family. The germline sequence comprises the V region sequence 1-18, the D region sequences 6-13, 1-1 OR 1-7, and the J region sequence JH4.

Figure 35 shows a comparison of predicted Fab "RD-B1" heavy chain amino acid sequence (residues 1-119 inclusive in Figure 5) with germline sequence from the VH2 family. The germline sequence comprises the V
region sequence 2-26, the D region sequence 3-22, and the J region sequence JH5.
Figure 36 shows a comparison of predicted Fab "58C" heavy chain amino acid sequence (residues 1-119 inclusive in Figure 7) with germline sequence from the VH3 family. The germline sequence comprises the V
region sequence 3-21, the D region sequence unknown, and the J region sequence JH4.
Figure 37 shows a comparison of predicted Fab "GP-A" heavy chain amino acid sequence (residues 1-124 inclusive in Figure 8) with germline sequence from the VH3 family. The germline sequence comprises the V
region sequence 3-30.3, the D region sequence 3-10, and the J region sequence JH3.
Figure 38 shows a comparison of predicted Fab "57D" heavy chain amino acid sequence (residues 1-117 inclusive in Figure 9) with germline sequence from the VH3 family. The germline sequence comprises the V
region sequence 3-53, the D region sequence 3-16, and the J region sequence unknown.
Figure 39 shows a comparison of predicted Fab "57E" heavy chain amino acid sequence (residues 1-118 inclusive in Figure 10) with germline sequence from the VH4 family. The germline sequence comprises the the V
region sequence 4-61, the D region sequence 7-27, and the J region sequence JH4.

Figure 40 shows a comparison of predicted Fab "67C" heavy chain amino acid sequence (residues 1-119 inclusive in Figure 12) with germline sequence from the VH2 family. The germline sequence comprises the V
region sequence 2-05, the D region sequence 5-18, and the J region sequence JH6.
Figure 41 shows a comparison of predicted Fab "BS-A", "BS-B", "57E" and "59-A2" light chain amino acid sequences (residues 1-111, 1-110, 1-112 and 1-110 inclusive in Figures 14, 15, 21 and 24, respectively) with germline sequence from the VX6 family. The germline sequence comprises the V region sequence 6a, and the J region sequences unknown or JL2 or JL3.
Figure 42 shows a comparison of predicted Fab "RD-B1" light chain amino acid sequence (residues 1-112 inclusive in Figure 16) with germline sequence from the VK2 family. The germline sequence comprises the V
region sequence A23, and the J region sequence JK2.
Figure 43 shows a comparison of predicted Fab "RD-A2", "IFN-A" and "67C" light chain amino acid sequences (residues 1-112, 1-110, 1-112 and 1-110 inclusive in Figures 17, 22, and 23, respectively) with germline sequence from the VK2 family. The germline sequence comprises the V region sequence A19, and the J
region sequences JK3, JK2 and JK2, respectively.
Figure 44 shows a comparison of predicted Fab "58C" light chain amino acid sequence (residues 1-108 inclusive in Figure 18) with germline sequence from the VX3 family. The germline sequence comprises the V
region sequence 3h, and the J region sequence unknown.

Figure 45 shows a comparison of predicted Fab "GP-A" light chain amino acid sequence (residues 1-106 inclusive in Figure 19) with germline sequence from the VX3 family. The germline sequence comprises the V
region sequence 2-19, and the J region sequence JL2 or JL3.
Figure 46 shows a comparison of predicted Fab "57D" light chain amino acid sequence (residues 1-108 inclusive in Figure 20) with germline sequence from the VK3 family. The germline sequence comprises the V
region sequence A27, and the J region sequences JK5.
Figure 47 shows a comparison of Fab classes.
Fab class comparison was done using GCG (Genetics Computer Group, 575 Science Drive, Madison, WI 53711) PileUp program for multiple sequence comparison analysis. The symbol (**) indicates that the closest matching diversity (D) region or joining region (J), although related to known germ line sequences, could not be determined. The symbol (*) indicates that variations in 1, 2 or 3 residues occur in comparison to the identified joining region.
Detailed Description of the Invention The present invention provides for agents which selectively bind ("selective binding agents") human gamma interferon-gamma protein (hIFNy).
Preferably, the agents are IFNy antagonists or inhibitors which inhibit partially or completely at least one activity of IFNy, such as binding of IFNy to its cognate receptor. In one embodiment, the fully human antibody fragments selectively binds IFNy such that it partially or completely blocks the binding of IFNy to its cognate receptor and partially or completely inhibits IFNy activity.
The term "selective binding agent" refers to a molecule which preferentially binds IFNy. A
selective binding agent may include a protein, peptide, nucleic acid, carbohydrate, lipid, or small molecular weight compound. In a preferred embodiment, a selective binding agent is an antibody, such as polyclonal antibodies, monoclonal antibodies (mAbs), chimeric antibodies, CDR-grafted antibodies, anti-idiotypic (anti-Id) antibodies to antibodies that can be labeled in soluble or bound form, as well as fragments, regions or derivatives thereof, provided by known techniques, including, but not limited to enzymatic cleavage, peptide synthesis or recombinant techniques. The anti-IFNy selective binding agents of the present invention are capable of binding portions of IFNy that inhibit the binding of IFNy to the IFNy-R
receptor.
The antibodies and antigen binding domains of the invention bind selectively to IFNy, that is they bind preferentially to IFNy with a greater binding affinity than to other antigens. The antibodies may bind selectively to human IFNy, but also bind detectably to non-human IFNy, such as murine IFNy.
Alternatively, the antibodies may bind selectively to non-human IFNy, but also bind detectably to human IFNy.
Alternatively, the antibodies may bind exclusively to human IFNy, with no detectable binding to non-human IFNy.
The term "monoclonal antibody" refers to an antibody obtained from a population of substantially homogeneous antibodies wherein each monoclonal antibody will typically recognize a single epitope on the antigen. The term "monoclonal" is not limited to any particular method for making the antibody. For example, monoclonal antibodies of the invention may be made by the hybridoma method as described in Kohler et al.; Nature, 256:495 (1975) or may be isolated from phage libraries using the techniques as described herein, for example.
The term "antigen binding domain" or "antigen binding region" refers to that portion of the selective binding agent (such as an antibody molecule) which contains the amino acid residues that interact with an antigen and confer on the binding agent its specificity and affinity for the antigen. Preferably, the antigen binding region will be of human origin. In other embodiments, the antigen binding region can be derived from other animal species, in particular rodents such as rabbit, rat or hamster.
The term "epitope" refers to that portion of any molecule capable of being recognized by and bound by a selective binding agent (such as an antibody) at one or more of the binding agent's antigen binding regions. Epitopes usually consist of chemically active surface groupings of molecules such as amino acids or sugar side chains and have specific three dimensional structural characteristics as well as specific charge characteristics. By "inhibiting and/or neutralizing epitope" is intended an epitope, which, when bound by a selective binding agent, results in loss of biological activity of the molecule or organism containing the epitope, in vivo, in vitro, or in situ, more preferably in vivo, including binding of IFNy to its receptor.
The term "light chain" when used in reference to an antibody refers to two distinct types, called kappa (k) of lambda (X) based on the amino acid sequence of the constant domains.
The term "heavy chain" when used in reference to an antibody refers to five distinct types, called alpha, delta, epsilon, gamma and mu, based on the amino acid sequence of the heavy chain constant domain.
These distinct types of heavy chains give rise to five classes of antibodies, IgA, IgD, IgE, IgG and IgM, respectively, including four subclasses of IgG, namely IgGi, IgG2, IgG3 and IgGe The term "variable region" or "variable domain" refers to a portion of the light and heavy chains, typically about the amino-teiwinal 120 to 130 amino acids in the heavy chain and about 100 to 110 amino acids in the light chain, which differ extensively in sequence among antibodies and are used in the binding and specificity of each particular antibody for its particular antigen. The variability in sequence is concentrated in those regions called complimentarily determining regions (CDRs) while the more highly conserved regions in the variable domain are called framework regions (FR). The CDRs of the light and heavy chains are responsible for the interaction of the antibody with antigen.
The term "constant region" or "constant domain" refers to a carboxy terminal portion of the light and heavy chain which is not directly involved in binding of the antibody to antigen but exhibits various effector function, such as interaction with the Fc receptor.
The term "human interferon-gamma" or "human interferon-gamma polypeptide" refers to the polypeptides comprising the amino acid sequences described in PCT Publication WO 83/04053, and related polypeptides. Related polypeptides include allelic variants; splice variants; fragments;
derivatives; substitution, deletion, and insertion variants; fusion polypeptides; and interspecies homologs. Also encompassed are soluble forms of IFNy which is sufficient to generate an immunological response. IFNy may be a mature polypeptide, as defined herein, and may or may not have an amino terminal methionine residue, depending upon the method by which it is prepared.
The term "fragment" when used in relation to IFNy or to a proteinaceous selective binding agent of IFNy refers to a peptide or polypeptide that comprises less than the full length amino acid sequence. Such a fragment may arise, for example, from a truncation at the amino terminus, a truncation at the carboxy terminus, and/or an internal deletion of a residue(s) from the amino acid sequence. Fragments may result from alternative ma splicing or from in vivo protease activity.
The term "variant" when used in relation to IFNy or to a proteinaceous selective binding agent of IFNy refers to a peptide or polypeptide comprising one or more amino acid sequence substitutions, deletions, and/or additions as compared to a native or unmodified sequence. For example, an IFNy variant may result from one or more changes to an amino acid sequence of native IFNy. Also by way of example, a variant of a selective binding agent of IFNy may result from one or more changes to an amino acid sequence of a native or previously unmodified selective binding agent.
Variants may be naturally occurring, such as allelic or splice variants, or may be artificially constructed.
Polypeptide variants may be prepared from the corresponding nucleic acid molecules encoding said variants.
The term "derivative" when used in relation to IFNy or to a proteinaceous selective binding agent of IFNy refers to a polypeptide or peptide, or a variant, fragment or derivative thereof, which has been chemically modified. Examples include covalent attachment of one or more polymers, such as water soluble polymers, N-linked, or 0-linked carbohydrates, sugars, phosphates, and/or other such molecules. The derivatives are modified in a manner that is different from naturally occurring or starting peptide or polypeptides, either in the type or location of the molecules attached. Derivatives further include deletion of one or more chemical groups which are naturally present on the peptide or polypeptide.
The term "fusion" when used in relation to IFNy or to a proteinaceous selective binding agent of IFNy refers to the joining of a peptide or polypeptide, or fragment, variant and/or derivative thereof, with a heterologous peptide or polypeptide.
The term "biologically active" when used in relation to IFNy or to a proteinaceous selective binding agent refers to a peptide or a polypeptide having at least one activity characteristic of IFNy or a selective binding agent. A selective binding agent of IFNy may have agonist, antagonist, or neutralizing or blocking activity with respect to at least one biological activity of IFNy.
The term "naturally occurring" when used in connection with biological materials such as nucleic acid molecules, polypeptides, host cells, and the like, refers to those which are found in nature and not manipulated by a human being.

The term "isolated" when used in relation to IFNy or to a proteinaceous selective binding agent of IFNy refers to a peptide or polypeptide that is free from at least one contaminating polypeptide that is found in its natural environment, and preferably substantially free from any other contaminating mammalian polypeptides which would interfere with its therapeutic or diagnostic use.
The term "mature" when used in relation to IFNy or to a proteinaceous selective binding agent of IFNy refers to a peptide or polypeptide lacking a leader sequence. The term may also include other modifications of a peptide or polypeptide such as proteolytic processing of the amino terminus (with or without a leader sequence) and/or the carboxy terminus, cleavage of a smaller polypeptide from a larger precursor, n-linked and/or o-linked glycosylation, and the like.
The terms "effective amount" and "therapeutically effective amount" when used in relation to a selective binding agent of IFNy refers to an amount of a selective binding agent that is useful or necessary to support an observable change in the level of one or more biological activities of IFNy.
Said change may be either an increase or decrease in the level of IFNy activity.
The term "conservative amino acid substitution" refers to a substitution of a native amino acid residue with a non-native residue such that there is little or no effect on the polarity or charge of the amino acid residue at that position. For example, a conservative substitution results from the replacement of a non-polar residue in a polypeptide with any other non-polar residue. Furthermore, any native residue in a polypeptide may also be substituted with alanine, as has been previously described for alanine scanning mutagenesis; Cunningham et al., Science, 244:1081-1085 (1989). Exemplary rules for conservative amino acid substitutions are set forth in Table I.
Table I
Conservative Amino Acid Substitutions Original Exemplary Preferred Residues Substitutions Substitutions ALA VAL,LEU,ILE VAL
ARG LYS,GLN,ASN LYS
ASN GLN,HIS,LYS,ARG GLN
ASP GLU GLU
CYS SER SER
GLN ASN ASN
GLU ASP ASP
GLY PRO,ALA ALA
HIS ASN,GLN,LYS,ARG ARG
ILE LEU,VAL,MET,ALA, LEU
PHE,NORLEUCINE
LEU NORLEUCINE,ILE, ILE
VAL,MET,ALA,PHE
LYS ARG,GLN,ASN ARG
MET LEU,PHE,ILE LEU
PHE LEU,VAL,ILE,ALA, LEU
TYR
PRO ALA ALA
SER THR THR
THR SER SER
TRP TYR, PEE TYR

TYR TRP,PHE,THR,SER PHE
VAL ILE,MET,LEU,PHE, LEU
ALA, NORLEUCINE
Conservative amino acid substitutions also encompass non-naturally occurring amino acid residues which are typically incorporated by chemical peptide synthesis rather than by synthesis in biological systems. These include peptidomimetics, and other reversed or inverted forms of amino acid moieties.
Conservative modifications to the amino acid sequence (and the corresponding modifications to the encoding nucleotides) are able to produce IFNy polypeptides (and proteinaceous selective binding agents thereof) having functional and chemical characteristics similar to those of naturally occurring IFNy or selective binding agents. In contrast, substantial modifications in the functional and/or chemical characteristics of IFNy (and protineaceous selective binding agents thereof) may be accomplished by selecting substitutions that differ significantly in their effect on maintaining (a) the structure of the molecular backbone in the area of the substitution, for example, as a sheet or helical conformation, (b) the charge or hydrophobicity of the molecule at the target site, or (c) the bulk of the side chain. Naturally occurring residues may be divided into groups based on common side chain properties:
1) Hydrophobic: norleucine, Met, Ala, Val, Leu, Ile;
2) Neutral hydrophilic: Cys, Ser, Thr;
3) Acidic: Asp, Glu;
4) Basic: Asn, Gln, His, Lys, Arg;

5) Residues that influence chain orientation:
Gly, Pro; and 6) Aromatic: Trp, Tyr, Phe.
Non-conservative substitutions may involve the exchange of a member of one of these classes for a member from another class.
The "identity or similarity" of two or more nucleic acid molecules and/or polypeptides provides a measure of the relatedness of two or more distinct sequences. The term "identity" refers to amino acids which are identical at corresponding positions in two distinct amino acid sequences. The term "similarity"
refers to amino acids which are either identical or are conservative substitutions as defined above at corresponding positions in two distinct amino acid sequences.
The extent of identity or similarity can be readily calculated by known methods, including but not limited to those described in Computational Molecular Biology, Lesk, A.M., ed., Oxford University Press, New York, 1988; Biocamputing: Informatics and Ganome Projects, Smith, D.W., ed., Academic Press, New York, 1993; Computer Analysis of Sequence Data, Part 1, Griffin, A.M., and Griffin, H.G., eds., Humana Press, New Jersey, 1994; Sequence Analysis in Molecular Biology, von Heinje, G., Academic Press, 1987; Sequence Analysis Primer, Gribskov, M. and Devereux, J., eds., M. Stockton Press, New York, 1991; and Carillo et al., SIAM J. Applied Math., 48:1073 (1988).
Preferred methods to determine identity and/or similarity are designed to give the largest match between the sequences tested. Methods to determine identity and similarity are codified in publicly available computer programs. Exemplary computer program methods to determine identity and similarity between two sequences include, but are not limited to, the GCG program package, including GAP;
Devereux et al., Nucleic Acids Research, 12:387 (1984);
Genetics Computer Group, University of Wisconsin, Madison, WI; BLASTP, BLASTN, and FASTA Altschul et al., J. Mol. Biol., 215:403-410 (1990). The BLAST X program is publicly available from the National Center for Biotechnology Information (NCBI) and other sources (BLAST Manual, Altschul et al. NCB NLM NIH Bethesda, MD). The well known Smith Waterman algorithm may also be used to determine identity.
'FM/ polvpaptides IFNy polypeptides, and fragments, variants and derivatives thereof, are used as target molecules for screening and identifying the selective binding agents of the invention. When it is desired to prepare antibodies as selective binding agents, IFNI, polypeptides are preferably immunogenic, that is they elicit an immune response when administered to an animal. Alternatively, when antibodies are prepared by in vitro techniques, IFNy polypeptides used as target molecules are capable of detectably binding an antibody or antigen binding domain.
IFNy polypeptides are prepared by biological or chemical methods. Biological methods such as expression of DNA sequences encoding recombinant 'FM, are known in the art; see e.g., Sambrook et al. Supra.
Chemical synthesis methods such as those set forth by Merrifield et al.,J. Am. Chem. Soc., 85:2149 (1963), Houghten et al., Proc Natl Acad. Sci. USA, 82:5132 (1985), and Stewart and Young, Solid phase peptide synthesis, Pierce Chemical Co., Rockford, IL (1984) may also be used to prepare IFN/ polypeptides of the invention. Such polypeptides may be synthesized with or without a methionine on the amino terminus.
Chemically synthesized 'FM, polypeptides, or fragments or variants thereof, may be oxidized using methods set forth in these references to form disulfide bridges.
IFNy polypeptides of the invention prepared by chemical synthesis will have at least one biological activity comparable to the corresponding 'FM polypeptides produced recombinantly or purified from natural sources.
IFNy polypeptides may be obtained by isolation from biological samples such as source tissues and/or fluids in which the IFNy polypeptides are naturally found. Sources for IFNy polypeptides may be human or non-human in origin. Isolation of naturally-occurring IFNy polypeptides can be accomplished using methods known in the art, such as separation by electrophoresis followed by electroelution, various types of chromatography (affinity, immunoaffinity, molecular sieve, and/or ion exchange), and/or high pressure liquid chromatography.
The presence of the IFNy polypeptide during purification may be monitored using, for example, an antibody prepared against recombinantly produced IFNy polypeptide or peptide fragments thereof.
Polypeptides of the invention include isolated IFNy polypeptides and polypeptides related thereto including fragments, variants, fusion polypeptides, and derivatives as defined hereinabove.
'FM fragments of the invention may result from truncations at the amino terminus (with or without a leader sequence), truncations at the carboxy terminus, and/or deletions internal to the polypeptide. Such IFNy polypeptides fragments may optionally comprise an amino terminal methionine residue. The polypeptides of the invention will be immunogenic in that they will be capable of eliciting an antibody response.
IFNy polypeptide variants of the invention include one or more amino acid substitutions, additions and/or deletions as compared to the native IFNy amino acid sequence. Amino acid substitutions may be conservative, as defined above, or non-conservative or any combination thereof. The variants may have additions of amino acid residues either at the carboxy terminus or at the amino terminus (where the amino terminus may or may not comprise a leader sequence).
Embodiments of the invention include IFNy glycosylation variants and cysteine variants. IFNy glycosylation variants include variants wherein the number and/or type of glycosylation sites has been altered compared to native IFNy polypeptide. In one embodiment, IFNy glycosylation variants comprise a greater or a lesser number of N-linked glycosylation sites compared to native IFNy.
Also provided for are IFNy glycoyslation variants comprising a rearrangement of N-linked carbohydrate chains wherein one or more N-linked glycosylation sites (typically those that are naturally occurring) are eliminated and one or more new N-linked sites are created. IFNy cysteine variants comprise a greater number or alternatively a lesser number of cysteine residues compared to native IFNy. In one embodiment, one or more cysteine residues are deleted or substituted with another amino acid (e.g., serine).
Cysteine variants of IFNy can improve the recovery of biologically active IFNy by aiding the refolding of IFNy into a biologically active conformation after isolation from a denatured state.
Preparing IFNy polypeptide variants is within the level of skill in the art. In one approach, one may introduce one or more amino acid substitutions, deletions and/or additions in native IFNy wherein the IFNy variant retains the native structure of IFNy and/or at least one of the biological activities. One approach is to compare sequences of IFNy polypeptides from a variety of different species in order to identify regions of relatively low and high identity and/or similarity. It is appreciated that those regions of an IFNy polypeptide having relatively low identity and/or similarity, are less likely to be essential for structure and activity and therefore may be more tolerant of amino acid alterations, especially those which are non-conservative. It is also appreciated that even in relatively conserved regions, one could introduce conservative amino acid substitutions while retaining activity.
In another approach, structure-function relationships can be used to identify residues in similar polypeptides that are important for activity or structure. For example, one may compare conserved amino acid residue among IFNy and other members of the tumor necrosis factor family for which structure-function analyses are available and, based on such a comparison, predict which amino acid residues in IFNT
are important for activity or structure. One skilled in the art may choose chemically similar amino acid substitutions for such predicted important amino acid residues of IFNy.

In yet another approach, an analysis of a secondary or tertiary structure of IFNy (either determined by x-ray diffraction of IFNy crystals or by structure prediction methods) can be undertaken to determine the location of specific amino acid residues in relation to actual or predicted structures within an IFNy polypeptide. Using this information, one can introduce amino acid changes in a manner that seeks to retain as much as possible the secondary and/or tertiary structure of an IFNy polypeptide.
In yet another approach, the effects of altering amino acids at specific positions may be tested experimentally by introducing amino acid substitutions and testing the altered IFNy polypeptides for biological activity using assays described herein.
Techniques such as alanine scanning mutagenesis (Cunningham et al., supra) are particularly suited for this approach. Many altered sequence may be conveniently tested by introducing many substitutions at various amino acid positions in IFNy and screening the population of altered polypeptides as part of a phage display library. Using this approach, those regions of an IFNy polypeptide that are essential for activity may be readily determined.
The above methods are useful for generating IFNy variants which retain the native structure. Thus, antibodies raised against each variants are likely to recognize a native structural determinant, or epitope, of IFNy and are also likely to bind to native IFNy.
However, in some cases is may be desirable to produce IFNy variants which do not retain native IFNy structure or are partially or completely unfilled. Antibodies raised against such proteins will recognize buried epitopes on IFNy.
The invention also provides for IFNy fusion polypeptides which comprise IFNy polypeptides, and fragments, variants, and derivatives thereof, fused to a heterologous peptide or protein. Heterologous peptides and proteins include, but are not limited to:
an epitope to allow for detection and/or isolation of a IFNy fusion polypeptide; a transmembrane receptor protein or a portion thereof, such as an extracellular domain, or a transmembrane and intracellular domain; a ligand or a portion thereof which binds to a transmembrane receptor protein; an enzyme or portion thereof which is catalytically active; a protein or peptide which promotes oligomerization, such as leucine zipper domain; and a protein or peptide which increases stability, such as an immunoglobulin constant region.
A IFNy polypeptide may be fused to itself or to a fragment, variant, or derivative thereof. Fusions may be made either at the amino terminus or at the carboxy terminus of a IFNy polypeptide, and may be direct with no linker or adapter molecule or may be through a linker or adapter molecule. A linker or adapter molecule may also be designed with a cleavage site for a DNA restriction endonuclease or for a protease to allow for separation of the fused moieties.
In a further embodiment of the invention, a IFNy polypeptide, fragment, variant and/or derivative is fused to an Fc region of human IgG. In one example, a human IgG hinge, ch2 and ch3 region may be fused at either the N-terminus or C-terminus of the IFNy polypeptides using methods known to the skilled artisan. In another example, a portion of a hinge regions and ch2 and ch3 regions may be fused. The IFNy Fc-fusion polypeptide so produced may be purified by use of a protein a affinity column. In addition, peptides and proteins fused to an fc region have been found to exhibit a substantially greater half-life in vivo than the unfused counterpart. Also, a fusion to an Fc region allows for dimerization/multimerization of the fusion polypeptide. The Fc region may be a naturally occurring Fc region, or may be altered to improve certain qualities, such as therapeutic qualities, circulation time, reduce aggregation, etc.
IFNy polypeptide derivatives are included in the scope of the present invention. Such derivatives are chemically modified IFNy polypeptide compositions in which IFNy polypeptide is linked to a polymer. The polymer selected is typically water soluble so that the protein to which it is attached does not precipitate in an aqueous environment, such as a physiological environment. The polymer may be of any molecular weight, and may be branched or unbranched. Included within the scope of IFNy polypeptide polymers is a mixture of polymers. Preferably, for therapeutic use of the end-product preparation, the polymer will be pharmaceutically acceptable.
The water soluble polymer or mixture thereof may be for example, polyethylene glycol (PEG), monomethoxy-polyethylene glycol, dextran (such as low molecular weight dextran, of, for example about 6 kD), cellulose, or other carbohydrate based polymers, poly-(N-vinyl pyrrolidone) polyethylene glycol, propylene glycol homopolymers, a polypropylene oxide/ethylene oxide co-polymer, polyoxyethylated polyols (e.g., glycerol) and polyvinyl alcohol.
A preferred water soluble polymer is polyethylene glycol. As used herein, polyethylene glycol is meant to encompass any of the forms of PEG

that have been used to derivatize other proteins, such as mono- (C1-C10) alkoxy-, or aryloxy-polyethylene glycol. Also encompassed by the invention are bifunctional PEG crosslinking molecules which may be used to prepare covalently attached IFNy multimers.
Methods for preparing chemically derivatized IFNy polypeptides are known in the art. By way of example, derivatization of IFNy,polypeptides with PEG
may be carried out using procedures described in Francis et al., Focus on Growth Factors, 3:4-10 (1992);
EP 0 154 316; and EP 0 401 384. In a preferred embodiment, an IFNy polypeptide derivative will have a single PEG moiety at the amino terminus; see U.S.
Patent No. 5,985,265.
IFNy polypeptide derivatives disclosed herein may exhibit an enhancement or reduction of at least one biological activity of IFNy compared to unmodified polypeptide, or may exhibit increased or decreased half-life or stability.
IFNI, selective binding agents IFNy polypeptides, and fragments, variants and derivatives thereof, may be used to identify selective binding agents of 'FM. As defined above, a selective binding agent of IFNy encompasses both proteinaceous and non-proteinaceous binding agents and, in one preferred embodiment of the invention, the selective binding agent is proteinaceous. In yet another preferred embodiment, the selective binding agent is an antibody or fragment thereof which binds IFNy, preferably human IFNy. The antibodies of the invention may be agonist antibodies, which enhance the level of at least one biological activity of IFNy; or antagonist antibodies, which decrease the level of at least one biological activity of IFNy. Antagonist antibodies of IFNy may also be referred to as inhibitory or neutralizing antibodies of IFNy.
Although such antibodies are preferred embodiments of the invention, it is understood that other proteinaceous selective binding agents which are agonists or antagonists of IFNy activity are also encompassed by the invention.
As described in the examples below, anti-IFNy antibodies and antigen binding domains which inhibit at least one activity of IFNy have been identified. Embodiments of the invention include antibodies comprising a heavy chain Fab sequence as shown in any of Figures 3-13 and further comprising a kappa or lambda light chain sequence. Light chain Fab sequences may be as shown in Figures 14-24. For example, "BS-A" antibody has light and heavy chain sequences in Figures 14 and 3, respectively; "BS-B"
antibody has light and heavy chains sequences of Figures 15 and 4, respectively; "RD-B1" antibody has light and heavy chain sequences of Figures 16 and 5, respectively; "RD-A2" antibody has light and heavy chain sequences of Figures 17 and 6, respectively;
"58C" antibody has light and heavy chain sequences of Figures 18 and 7, respectively; "GP-A" antibody has light and heavy chain sequences of Figures 19 and 8, respectively; "57D" antibody has light and heavy chain sequences of Figures 20 and 9, respectively; "57E"
antibody has light and heavy chain sequences of Figures 21 and 10, respectively; "IFN-A" antibody has light and heavy chain sequences of Figures 22 and 11, respectively; "67C" antibody has light and heavy chain sequences of Figures 23 and 12, respectively; and "59-A2" antibody has light and heavy chain sequences of Figures 24 and 13, respectively. The antibodies of the invention further comprise a human Fc region from any isotype, either IgG, IgM, IgA, IgE, or IgD.
Preferably, the Fc region is from human IgG, such as IgG1, IgG2, IgG3, or IgG4.
The invention also provides for antibodies or antigen binding domains which comprise fragments, variants, or derivatives of the Fab sequences disclosed herein. Fragments include variable domains of either the light or heavy chain Fab sequences which are typically joined to light or heavy constant domains.
Variants include antibodies comprising light chain Fab sequences which are at least about 80%, 85%, 90%, 95%, 98% or 99% identical or similar to the Fab sequences, or the corresponding variable domains, in any one of Figures 14-24, or antibodies comprising heavy chain Fab sequences, or the corresponding variable domains, which are at least about 80%, 85%, 90%, 95%, 98% or 99%
identical or similar to the Fab sequences in any one of Figures 3-13. The antibodies may be typically associated with constant regions of the heavy and light chains to form full-length antibodies.
Antibodies and antigen binding domains, and fragments, variants and derivatives thereof, of the invention will retain the ability to bind selectively to an IFN7 polypeptide, preferably to a human IFN'y polypeptide. In one embodiment, an antibody will bind an 'FM polypeptide with a dissociation constant (KD) of about 1nM or less, or alternatively 0.1nM or less, or alternatively 10pM or less or alternatively less than 10pM.
Antibodies of the invention include polyclonal monospecific polyclonal, monoclonal, recombinant, chimeric, humanized, fully human, single chain and/or bispecific antibodies. Antibody fragments include those portions of an anti- IFNy antibody which bind to an epitope on an IFNy polypeptide. Examples of such fragments include Fab F(ab'), F(ab)', Fv, and sFv fragments. The antibodies may be generated by enzymatic cleavage of full-length antibodies or by recombinant DNA techniques, such as expression of recombinant plasmids containing nucleic acid sequences encoding antibody variable regions.
Polyclonal antibodies are heterogeneous populations of antibody molecules derived from the sera of animals immunized with an antigen. An antigen is a molecule or a portion of a molecule capable of being bound by an antibody which is additionally capable of inducing an animal to produce antibody capable of binding to an epitope of that antigen. An antigen can have one or more epitope. The specific reaction referred to above is meant to indicate that the antigen will react, in a highly selective manner, with its corresponding antibody and not with the multitude of other antibodies which can be evoked by other antigens.
Polyclonal antibodies directed toward an IFNy polypeptide generally are raised in animals (e.g., rabbits or mice) by multiple subcutaneous or intraperitoneal injections of IFNy and an adjuvant. In accordance with the invention, it may be useful to conjugate an IFNy polypeptide, or a variant, fragment, or derivative thereof to a carrier protein that is immunogenic in the species to be immunized, such as keyhole limpet heocyanin, serum, albumin, bovine thyroglobulin, or soybean trypsin inhibitor. Also, aggregating agents such as alum are used to enhance the immune response. After immunization, the animals are bled and the serum is assayed for anti- IFNy antibody titer.
Monoclonal antibodies (mAbs) contain a substantially homogeneous population of antibodies specific to antigens, which population contains substantially similar epitope binding sites. Such antibodies may be of any immunoglobulin class including IgG, IgM, IgE, IgA, IgD and any, subclass thereof. A
hybridoma producing a monoclonal antibody of the present invention may be cultivated in vitro, in situ, or in vivo. Production of high titers in vivo or in situ is a preferred method of production.
Monoclonal antibodies directed toward IFNy are produced using any method which provides for the production of antibody molecules by continuous cell lines in cultul-e.
Examples of suitable methods for preparing monoclonal antibodies include hybridoma methods of Kohler et al., Nature, 256:495-497 (1975), and the human B-cell hybridoma method, Kozbor, J. Immunol,. 133:3001 (1984);
Brodeur et al., Monoclonal Antibody Production Techniques and Applications, pp. 51-63 (Marcel Dekker, Inc., New York, 1987); and Harlow and Lane, Antibodies:
A Laboratory Manual, Cold Spring Harbor Laboratory (1988).
Preferred anti-IFNy selective binding agents include monoclonal antibodies which will inhibit partially or completely the binding of human IFNy to its cognate receptor, hIFNy-R, or an antibody having substantially the same Specific binding characteristics, as well as fragments and regions thereof. Preferred methods for determining monoclonal antibody specificity and affinity by competitive inhibition can be found in Harlow et al., Antibodies: A
=

Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., 1988); Colligan et al., eds., Current Protocols in Immunology, Greene Publishing Assoc. and Wiley Interscience, N.Y., (1992, 1993); and Muller, Meth. Enzymol., 92:589-601 (1983).
Also provided by the invention are hybridoma cell lines which produce monoclonal antibodies reactive with IFNy polypeptides.
Chimeric antibodies are molecules in which different portions are derived from different animal species, such as those having a variable region derived from a murine monoclonal antibody and a human immunoglobulin constant region. Chimeric antibodies are primarily used to reduce immunogenicity in application and to increase yields in production, for example, where murine monoclonal antibodies have higher yields from hybridomas but higher immunogenicity in humans, such that human/murine chimeric monoclonal antibodies are used.
Chimeric antibodies and methods for their production are known in the art. Cabilly et a/., Proc.
Natl. Acad. Sci. USA, 81:3273-3277 (1984); Morrison et al., Proc. Natl. Acad. Sci. USA, 81:6851-6855 (1984);
Boulianne et al., Nature, 312:643-646 (1984); Neuberger et a/., Nature, 314:268-270 (1985); Liu et al., Proc.
Natl. Acad. Sci. USA, 84:3439-3443 (1987); and Harlow and Lane Antibodies: A Laboratory Manual, Cold Spring Harbor Laboratory (1988).
For example, chimeric monoclonal antibodies of the invention may be used as a therapeutic. In such a chimeric antibody, a portion of the heavy and/or light chain is identical with or homologous to corresponding sequence in antibodies derived from a particular species or belonging to one particular antibody class or subclass, while the remainder of the chain(s) is identical with or homologous to corresponding sequence in antibodies derived from another species or belonging to another antibody class or subclass, as well as fragments of such antibodies, so long as they exhibit the desired biological activity; see, e.g., U.S. Patent No. 4,816,567 and Morrison et al., Proc. Natl. Acad. Sci., 81:6851-6855 (1985).
As used herein, the term "chimeric antibody"
includes monovalent, divalent or polyvalent immunoglobulins. A monovalent chimeric antibody is a dimer (HL) formed by a chimeric H chain associated through disulfide bridges with a chimeric L chain. A
divalent chimeric antibody is tetramer (H.L2) formed by two HL dimers associated through at least one disulfide bridge. A polyvalent chimeric antibody can also be produced, for example, by employing a C. region that aggregates (e.g., from an IgM H chain, or chain).
Murine and chimeric antibodies, fragments and regions of the present invention may comprise individual heavy (H) and/or light (L) immunoglobulin chains. A chimeric H chain comprises an antigen binding region derived from the H chain of a non-human antibody specific for IFNy, which is linked to at least a portion of a human H chain C region (CO, such as CH, or CH,.
A chimeric L chain according to the present invention comprises an antigen binding region derived from the L chain of a non-human antibody specific for IFNy, linked to at least a portion of a human L chain C
region (CL).
Selective binding agents, such as antibodies, fragments, or derivatives, having chimeric H chains and L chains of the same or different variable region binding specificity, can also be prepared by appropriate association of the individual polypeptide chains, according to known method steps; see, e.g., Ausubel et al., eds. Current Protocols in Molecular Biology, Wiley Interscience, N.Y. (1993) and Harlow et al., Antibodies: A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.
(1988). With this approach, hosts expressing chimeric H chains (or their derivatives) are separately cultured from hosts expressing chimeric L chains (or their derivatives), and the immunoglobulin chains are separately recovered and then associated. Alternatively, the hosts can be co-cultured and the chains allowed to associate spontaneously in the culture medium, followed by recovery of the assembled immunoglobulin, fragment or derivative.
As an example, the antigen binding region of the selective binding agent (such as a chimeric antibody) of the present invention is preferably derived from a non-human antibody specific for human IFNy. Preferred sources for the DNA encoding such a non-human antibody include cell lines which produce antibodies, such as hybrid cell lines commonly known as hybridomas.
The invention also provides for fragments, variants and derivatives, and fusions of anti-IFNy antibodies, wherein the terms "fragments", "variants", "derivatives" and "fusions" are defined herein. The _invention encompasses fragments, variants, derivatives, and fusions of anti-IFNy antibodies which are functionally similar to the unmodified anti-IFNy antibody, that is, they retain at least one of the activities of the unmodified antibody. In addition to the modifications set forth above, also included is the addition of genetic sequences coding for cytotoxic proteins such as plant and bacterial toxins. The fragments, variants, derivatives and fusions of anti-IFNy antibodies can be produced from any of the hosts of this invention.
Suitable fragments include, for example, Fab, Fab', F(ab')2, Fv and scFv. These fragments lack the Fc fragment of an intact antibody, clear more rapidly from the circulation, and can have less non-specific tissue binding than an intact antibody; Wahl et al., J.
Nucl. Med., 24:316-325 (1983). These fragments are produced from intact antibodies using methods well known in the art, for example by proteolytic cleavage with enzymes such as papain (to produce Fab frayments) or pepsin (to produce F(ab')2 fragments). The identification of these antigen binding regions and/or epitopes recognized by monoclonal antibodies of the present invention provides the information necessary to generate additional monoclonal antibodies with similar binding characteristics and therapeutic or diagnostic utility that parallel the embodiments of this application.
Variants of selective binding agents are also provided. In one embodiment, variants of antibodies and antigen binding domains comprise changes in light and/or heavy chain amino acid sequences that are naturally occurring or are introduced by in vitro engineering of native sequences using recombinant DNA
techniques. Naturally occurring variants include "somatic" variants which are generated in vivo in the corresponding germ line nucleotide sequences during the generation of an antibody response to a foreign antigen. Variants encoded by somatic mutations in germline variable light and heavy chain sequences which generate the exemplary Fabs of the present invention in sequences are shown in Figures 33 and 41 for Fab "BS-A", Figures 34 and 41 for Fab "BS-B", Figures 35 and 42 for Fab "RD-B1", Figures 35 and 42 for Fab "RD-B1", Figures 33 and 43 for Fab "RD-A2", Figures 36 and 44 for Fab "58C", Figures 37 and 45 for Fab "GP-A", Figures 38 and 46 for Fab "57D", Figures 39 and 41 for Fab "57E", Figures 33 and 43 for Fab "IFN-A", Figures 40 and 43 for Fab "67C", and Figures 34 and 41 for Fab "59-A2".
Variants of anti-IFNy antibodies and antigen binding domains can also be prepared by mutagenesis techniques known in the art. In one example, amino acid changes may be introduced at random throughout an antibody coding region and the resulting variants may be screened for a desired activity, such as binding affinity for IFNy. Alternatively, amino acid changes may be introduced in selected regions of an IFNy antibody, such as in the light and/or heavy chain CDRs, and framework regions, and the resulting antibodies may be screened for binding to IFNy or some other activity.
Amino acid changes encompass one or more amino acid substitutions in a CDR, ranging from a single amino acid difference to the introduction of all possible permutations of amino acids within a given CDR, such as CDR3. In another method, the contribution of each residue within a CDR to IFNy binding may be assessed by substituting at least one residue within the CDR with alanine; Lewis et al., MO1. ImmunoI., 32:1065-1072 (1995). Residues which are not optimal for binding to IFNy may then be changed in order to determine a more optimum sequence. Also encompassed are variants generated by insertion of amino acids to increase the size of a CDR, such as CDR3. For example, most light chain CDR3 sequences are nine amino acids in length.
Light chain CDR3 sequences in an antibody which are shorter than nine residues may be optimized for binding to IFN7 by insertion of appropriate amino acids to increase the length of the CDR.
In one embodiment, antibody or antigen binding domain variants comprise one or more amino acid changes in one or more of the heavy or light chain CDR1, CDR2 or CDR3 and optionally one or more of the heavy or light chain framework regions FR1, FR2 or FR3.
Amino acid changes comprise substitutions, deletions and/or insertions of amino acid residues. Exemplary variants include an "BS-A" heavy chain variable region variant with one or more amino acid changes in the sequences GYYWS (SEQ ID NO:34); EINHSGSTNYNPSLKS (SEQ
ID NO:44); or GRARNWRSRFDY (SEQ ID NO:54), or an "BS-A"
light chain variable region variant with one or more amino acid changes in the sequences TGSSGSIASHYVQ (SEQ
ID NO:01); EDKERPS (SEQ ID NO:12); or QSYDSSNQWV (SEQ
ID NO:23). The aforementioned "BS-A" heavy and light chain variable region variants may further comprise one or more amino acid changes in the framework regions.
In one example, one or more amino acid changes may be introduced to substitute a somatically mutated framework residue with the germline residue at that position. When the aforementioned amino acid changes are substitutions, the changes may be conservative or non-conservative substitutions.
Variants may also be prepared by "chain shuffling" of either light or heavy chains; Marks et al.
Biotechnology, 10:779-783 (1992). Typically, a single light (or heavy) chain is combined with a library having a repertoire of heavy (or light) chains and the resulting population is screened for a desired activity, such as binding to IFNy. This technique permits screening of a greater sample of different heavy (or light) chains in combination with a single light (or heavy) chain than is possible with libraries comprising repertoires of both heavy and light chains.
The selective binding agents of the invention can be bispecific. Bispecific selective binding agents of this invention can be of several configurations.
For example, bispecific antibodies resemble single antibodies (or antibody fragments) but have two different antigen binding sites (variable regions).
Bispecific antibodies can be produced by chemical techniques; see e.g., Kranz et al., Proc. Natl. Acad.
Sci. USA, 78:5807 (1981); by "polydoma" techniques;
U.S. Pat. No. 4,474,893; or by recombinant DNA
techniques.
The selective binding agents of the invention may also be heteroantibodies. Heteroantibodies are two or more antibodies, or antibody binding fragments (Fab) linked together, each antibody or fragment having a different specificity.
The invention also relates to "humanized"
antibodies. Methods for humanizing non-human antibodies are well known in the art. Generally, a humanized antibody has one or more amino acid residues introduced into a human antibody from a source which is non-human. In general, non-human residues will be present in CDRs. Humanization can be performed following methods known in the art; Jones et al., Nature 321:522-525 (1986); Riechmann et a/., Nature, 332:323-327 (1988); Verhoeyen et al., Science, 239:1534-1536 (1988), by substituting rodent complementarily-determining regions (CDRs) for the corresponding regions of a human antibody.

The selective binding agents of the invention, including chimeric, CDR-grafted, and humanized antibodies can be produced by recombinant methods known in the art. Nucleic acids encoding the antibodies are introduced into host cells and expressed using materials and procedures described herein and known in the art. In a preferred embodiment, the antibodies are produced in mammalian host cells, such as CHO cells. Fully human antibodies may be produced by expression of recombinant DNA transfected into host cells or by expression in hybridoma cells as described above.
Techniques for creating recombinant DNA
versions of the antigen-binding regions of antibody molecules which bypass the generation of monoclonal antibodies are encompassed within the practice of this invention. To do so, antibody-specific messenger RNA
molecules are extracted from immune system cells taken from an immunized animal, and transcribed into complementary DNA (cDNA). The cDNA is then cloned into a bacterial expression system. One example of such a technique suitable for the practice of this invention uses a filamentous bacteriophage M13 derived phagemid vector system having a leader sequence that causes the expressed Fab protein to migrate to the periplasmic space (between the bacterial cell membrane and the cell wall) or to be secreted. One can rapidly generate and screen great numbers of functional Fab fragments for those which bind the antigen. Such IFNy selective binding agents (Fab fragments with specificity for an IFNy polypeptide) are specifically encompassed within the term "antibody" as it is defined, discussed, and claimed herein.
Also within the scope of the invention are techniques developed for the production of chimeric antibodies by splicing the genes from a mouse antibody molecule of appropriate antigen-specificity together with genes from a human antibody molecule of appropriate biological activity, such as the ability to activate human complement and mediate ADCC; Morrison et al., Proc. Natl. Acad. Sci., 81:6851 (1984); Neuberger et a/., Nature, 312:604 (1984). One example is the replacement of a Fc region with that of a different isotype. Selective binding agents such as antibodies produced by this technique are within the scope of the invention.
In a preferred embodiment of the invention, the anti-IFM antibodies are fully human antibodies.
Thus encompassed by the invention are antibodies which bind IFNly polypeptides and are encoded by nucleic acid sequences which are naturally occurring somatic variants of human germline immunoglobulin nucleic acid sequence, and fragments, synthetic variants, derivatives and fusions thereof. Such antibodies may be produced by any method known in the art. Exemplary methods include immunization with a IFNy antigen (any IFNy polypeptide capable of elicing an immune response, and optionally conjugated to a carrier) of transgenic animals (e.g., mice) that are capable of producing a repertoire of human antibodies in the absence of endogenous immunoglobulin production; see, e.g., Jakobovits et al., Proc. Natl. Acad. Sci., 90:2551-2555 (1993); Jakobovits et al., Nature, 362:255-258 (1993);
Bruggermann et al., Year in Immunol., 7:33 (1993).
Alternatively, human antibodies may be generated through the in vitro screening of phage display antibody libraries; see e.g., Hoogenboom et a/., J. Mal. Biol., 227:381 (1991); Marks et al., J.
Mbl. Biol., 222:581 (1991).
Various antibody-containing phage display libraries have been described and may be readily prepared by one skilled in the art. Libraries may contain a diversity of human antibody sequences, such as human Fab, Fv, and scFv fragments, that may be screened against an appropriate target. Example 2 describes the screening of a Fab phage library against IFNy to identify those molecules which selectively bind IFNy. It will be appreciated that phage display libraries may comprise peptides or proteins other than antibodies which may be screened to identify selective binding agents of IFNy.
An anti-idiotypic (anti-Id) antibody is an antibody which recognizes unique determinants generally associated with the antigen-binding site of an antibody. An Id antibody can be prepared by immunizing an animal of the same species and genetic type (e.g., mouse strain) as the source of the monoclonal antibody with the monoclonal antibody to which an anti-Id is being prepared. The immunized animal will recognize and respond to the idiotypic determinants of the immunizing antibody by producing an antibody to these idiotypic determinants (the anti-Id antibody); see, e.g., U.S. Pat. No. 4,699,880.
The anti-Id antibody may also be used as an "immunogen" to induce an immune response in yet another animal, producing a so-called anti-anti-Id antibody. The anti-anti-Id may be epitopically identical to the original monoclonal antibody which induced the anti-Id. Thus, by using antibodies to the idiotypic determinants of a mAb, it is possible to identify other clones expressing antibodies of identical specificity.
Production of selective binding agents of IFNy When the selective binding agent of IFNy to be prepared is a proteinaceous selective binding agent, such as an antibody or an antigen binding domain, various biological or chemical methods for producing said agent are available.
Biological methods are preferable for producing sufficient quantities of a selective binding agent for therapeutic use. Standard recombinant dna techniques are particularly useful for the production of antibodies and antigen binding domains of the invention. Exemplary expression vectors, host cells and methods for recovery of the expressed product are described below.
A nucleic acid molecule encoding an IFNy such that amplification of the gene and/or expression of the gene can occur). A nucleic acid molecule encoding an anti-IFNy antibody may be amplified/expressed in prokaryotic, yeast, insect (baculovirus systems) and/or eukaryotic host cells.
Selection of the host cell will depend in part on whether an anti-IFNy antibody is to be post-transitionally modified (e.g., glycosylated and/or phosphorylated). If so, yeast, insect, or mammalian host cells are preferable. For a review of expression Typically, expression vectors used in any host cells will contain one or more of the following components: a promoter, one or more enhancer sequences, an origin of replication, a transcriptional termination sequence, a complete intron sequence containing a donor and acceptor splice site, a leader sequence for secretion, a ribosome binding site, a polyadenylation sequence, a polylinker region for inserting the nucleic acid encoding the polypeptide to be expressed, and a selectable marker element. Each of these sequences is discussed in more detail below.
The vector components may be homologous (i.e., from the same species and/or strain as the host cell), heterologous (i.e., from a species other than the host cell species or strain), hybrid (i.e., a combination of different sequences from more than one source), synthetic, or native sequences which normally function to regulate immunoglobulin expression. As such, a source of vector components may be any prokaryotic or eukaryotic organism, any vertebrate or invertebrate organism, or any plant, provided that the components are functional in, and can be activated by, the host cell machinery.
An origin of replication is selected based upon the type of host cell being used for expression.
For example, the origin of replication from the plasmid pbr322 (product no. 303-3s, New England Biolabs, Beverly, MA) is suitable for most gram-negative bacteria while various origins from SV40, polyoma, adenovirus, vesicular stomatitus virus (VSV) or papillomaviruses (such as HPV or BPV) are useful for cloning vectors in mammalian cells. Generally, the origin of replication component is not needed for mammalian expression vectors (for example, the SV40 origin is often used only because it contains the early promoter).
A transcription termination sequence is typically located 3' of the end of a polypeptide coding regions and serves to terminate transcription.
Usually, a transcription termination sequence in prokaryotic cells is a G-C rich fragment followed by a poly T sequence. While the sequence is easily cloned from a library or even purchased commercially as part of a vector, it can also be readily synthesized using methods for nucleic acid synthesis such as those described above.
A selectable marker gene element encodes a protein necessary for the survival and growth of a host cell grown in a selective culture medium. Typical selection marker genes encode proteins that (a) confer resistance to antibiotics or other toxins, e.g., ampicillin, tetracycline, or kanamycin for prokaryotic host cells, (b) complement auxotrophic deficiencies of the cell; or (c) supply critical nutrients not available from complex media. Preferred selectable markers are the kanamycin resistance gene, the ampicillin resistance gene, and the tetracycline resistance gene. A neomycin resistance gene may also be used for selection in prokaryotic and eukaryotic host cells.
Other selection genes may be used to amplify the gene which will be expressed. Amplification is the process wherein genes which are in greater demand for the production of a protein critical for growth are reiterated in tandem within the chromosomes of successive generations of recombinant cells. Examples of suitable selectable markers for mammalian cells include dihydrofolate reductase (DHFR) and thymidine kinase. The mammalian cell transformants are placed under selection pressure which only the transformants are uniquely adapted to survive by virtue of the marker present in the vector. Selection pressure is imposed by culturing the transformed cells under conditions in which the concentration of selection agent in the medium is successively changed, thereby leading to amplification of both the selection gene and the DNA

that encodes an anti-IFNy antibody. As a result, increased quantities of an antibody are synthesized from the amplified DNA.
A ribosome binding site is usually necessary for translation initiation of mrna and is characterized by a Shine-Dalgarno sequence (prokaryotes) or a Kozak sequence (eukaryotes). The element is typically located 3' to the promoter and 5' to the coding sequence of the polypeptide to be expressed. The Shine-Dalgarno sequence is varied but is typically a polypurine (i.e., having a high A-G content). Many Shine-Dalgarno sequences have been identified, each of which can be readily synthesized using methods set forth above and used in a prokaryotic vector.
A leader, or signal, sequence is used to direct secretion of a polypeptide. A signal sequence may be positioned within or directly at the 5' end of a polypeptide coding region. Many signal sequences have been identified and may be selected based upon the host cell used for expression. In the present invention, a signal sequence may be homologous (naturally occurring) or heterologous to a nucleic acid sequence encoding an anti-IFNy antibody or antigen binding domain. A
heterologous signal sequence selected should be one that is recognized and processed, i.e., cleaved, by a signal peptidase, by the host cell. For prokaryotic host cells that do not recognize and process a native immunoglobulin signal sequence, the signal sequence is substituted by a prokaryotic signal sequence selected, for example, from the group of the alkaline phosphatase, penicillinase, or heat-stable enterotoxin II leaders. For yeast secretion, a native immunoglobulin signal sequence may be substituted by the yeast invertase, alpha factor, or acid phosphatase leaders. In mammalian cell expression the native signal sequence is satisfactory, although other mammalian signal sequences may be suitable.
In most cases, secretion of an anti-IFNT
antibody or antigen binding domain from a host cell will result in the removal of the signal peptide from the antibody. Thus the mature antibody will lack any leader or signal sequence.
In some cases, such as where glycosylation is desired in a eukaryotic host cell expression system, one may manipulate the various presequences to improve glycosylation or yield. For example, one may alter the peptidase cleavage site of a particular signal peptide, or add prosequences, which also may affect glycosylation. The final protein product may have, in the -1 position (relative to the first amino acid of the mature protein) one or more additional amino acids incident to expression, which may not have been totally removed. For example, the final protein product may have one or two amino acid found in the peptidase cleavage site, attached to the N-terminus.
Alternatively, use of some enzyme cleavage sites may result in a slightly truncated form of the desired IFNy polypeptide, if the enzyme cuts at such area within the mature polypeptide.
The expression vectors of the present invention will typically contain a promoter that is recognized by the host organism and operably linked to a nucleic acid molecule encoding an anti-IFNT antibody or antigen binding domain. Either a native or heterologous promoter may be used depending the host cell used for expression and the yield of protein desired.
Promoters suitable for use with prokaryotic hosts include the beta-lactamase and lactose promoter systems; alkaline phosphatase, a tryptophan (trp) promoter system; and hybrid promoters such as the tac promoter. Other known bacterial promoters are also suitable. Their sequences have been published, thereby enabling one skilled in the art to ligate them to the desired DNA sequence(s), using linkers or adapters as needed to supply any required restriction sites.
Suitable promoters for use with yeast hosts are also well known in the art. Yeast enhancers are advantageously used with yeast promoters. Suitable promoters for use with mammalian host cells are well known and include those obtained from the genomes of viruses such as polyoma virus, fowlpox virus, adenovirus (such as Adenovirus 2), bovine papilloma virus, avian sarcoma virus, cytomegalovirus, a retrovirus, hepatitis-B virus and most preferably Simian Virus 40 (SV40). Other suitable mammalian promoters include heterologous mammalian promoters, e.g., heat-shock promoters and the actin promoter.
Additional promoters which may be used for expressing the selective binding agents of the invention include, but are not limited to: the SV40 early promoter region; Bernoist and Chambon, Nature, 290:304-310 (1981), the CMV promoter, the promoter contained in the 3' long terminal repeat of Rous sarcoma virus; Yamamoto, et al., Cell, 22:787-797 (1980), the herpes thymidine kinase promoter; Wagner et al., Proc. Natl. Acad. Sci. U.S.A., 78:144-1445 (1981), the regulatory sequences of the metallothionine gene;
Brinster et al., Nature, 296:39-42 (1982), prokaryotic expression vectors such as the beta -lactamase promoter; Villa-Kamaroff, et al., Proc. Natl. Acad.
Sci. U.S.A., 75:3727-3731 (1978), or the tac promoter;
DeBoer, et al., Proc. Natl. Acad. Sci. U.S.A., 80:21-25 (1983).
Also of interest are the following animal transcriptional control regions, which exhibit tissue specificity and have been utilized in transgenic animals: the elastase I gene control region which is active in pancreatic acinar cells; Swift et al., Cell, 38:639-646, (1984); Ornitz et al., Cold Spring Harbor Symp. Quant. Biol. 50:399-409 (1986); MacDonald, Ilepatology, 7:425-515 (1987), the insulin gene control region which is active in pancreatic beta cells;
Hanahan, Nature, 315:115-122 (1985), the immunoglobulin gene control region which is active in lymphoid cells;
Grosschedl et al., Cell, 38:647-658 (1984); Adames et al., Nature, 318:533-538 (1985); Alexander et al., Mbl.
Cell. Biol., 7:1436-1444 (1987), the mouse mammary tumor virus control region which is active in testicular, breast, lymphoid and mast cells; Leder et al., Cell, 45:485-495 (1986), the albumin gene control region which is active in liver; Pinkert et al., Genes and Devel., 1:268-276 (1987), the alphafetoprotein gene control region which is active in liver; Krumlauf et al., Mbl. Cell. Biol., 5:1639-1648 (1985); Hammer et al., Science, 235:53-58 (1987), the alpha 1-antitrypsin gene control region which is active in the liver;
Kelsey et al., Genes and Devel., 1:161-171 (1987); the beta-globin gene control region which is active in myeloid cells; Mogram et al., Nature, 315:338-340 (1985); Kollias et al., Cell, 46:89-94 (1986), the myelin basic protein gene control region which is active in oligodendrocyte cells in the brain; Readhead et al., Cell, 48:703-712 (1987), the myosin light chain-2 gene control region which is active in skeletal muscle; Sani, Nature, 314:283-286 (1985), and the gonadotropic releasing hormone gene control region which is active in the hypothalamus; Mason et al., Science, 234:1372-1378 (1986).
An enhancer sequence may be inserted into the vector to increase transcription in eucaryotic host cells. Several enhancer sequences available from mammalian genes are known (e.g., globin, elastase, albumin, alpha-feto-protein and insulin). Typically, however, an enhancer from a virus will be used. The SV40 enhancer, the cytomegalovirus early promoter enhancer, the polyoma enhancer, and adenovirus enhancers are exemplary enhancing elements for the activation of eukaryotic promoters. While an enhancer may be spliced into the vector at a position 5' or 3' to the polypeptide coding region, it is typically located at a site 5' from the promoter.
Preferred vectors for practicing this invention are those which are compatible with bacterial, insect, and mammalian host cells. Such vectors include, inter alia, pCRII, pCR3, and pcDNA3.1 (Invitrogen Company, San Diego, CA), pBSII (Stratagene Company, La Jolla, CA), pET15 (Novagen, Madison, WI), pGEX (Pharmacia Biotech, Piscataway, NJ), pEGFP-N2 (Clontech, Palo Alto, CA), pETL (BlueBacII; Invitrogen Carlsbad, CA), pDSR-alpha (PCT Publication No.
W090/14363) and pFastBacDual (Gibco/BRL, Grand Island, NY).
Additional possible vectors include, but are not limited to, cosmids, plasmids or modified viruses, but the vector system must be compatible with the selected host cell. Such vectors include, but are not limited to plasmids such as Bluescript plasmid derivatives (a high copy number Co1E1-based phagemid, Stratagene Cloning Systems Inc., La Jolla CA), PCR
cloning plasmids designed for cloning Taq-amplified PCR

products (e.g., TOPOI" TA Cloning Kit, PCR2.1 plasmid derivatives, Invitrogen, Carlsbad, CA), and mammalian, yeast or virus vectors such as a baculovirus expression system (pBacPAK plasmid derivatives, Clontech, Palo Alto, CA). The recombinant molecules can be introduced into host cells via transformation, transfection, infection, electroporation, or other known techniques.
Host cells of the invention may be prokaryotic host cells (such as E. coli) or eukaryotic host cells (such as a yeast cell, an insect cell, or a vertebrate cell). The host cell, when cultured under appropriate conditions, expresses an antibody or antigen binding domain of the invention which can subsequently be collected from the culture medium (if the host cell secretes it into the medium) or directly from the host cell producing it (if it is not secreted). Selection of an appropriate host cell will depend upon various factors, such as desired expression levels, polypeptide modifications that are desirable or necessary for activity, such as glycosylation or phosphorylation, and ease of folding into a biologically active molecule.
A number of suitable host cells are known in the art and many are available from the American Type Culture Collection (ATCC), Manassas, VA. Examples include mammalian cells, such as Chinese hamster ovary cells (CHO) (ATCC No. CCL61) CHO DHFR-cells; Urlaub et al., Proc. Natl. Acad. Sci. USA, 97:4216-4220 (1980), human embryonic kidney (HEK) 293 or 293T cells (ATCC
No. CRL1573), or 3T3 cells (ATCC No. CCL92). The selection of suitable mammalian host cells and methods for transformation, culture, amplification, screening and product production and purification are known in the art. Other suitable mammalian cell lines, are the monkey COS-1 (ATCC No. CRL1650) and COS-7 cell lines (ATCC No. CRL1651), and the CV-1 cell line (ATCC No.
CCL70). Further exemplary mammalian host cells include primate cell lines and rodent cell lines, including transformed cell lines. Normal diploid cells, cell strains derived from in vitro culture of primary tissue, as well as primary explants, are also suitable.

Candidate cells may be genotypically deficient in the selection gene, or may contain a dominantly acting selection gene. Other suitable mammalian cell lines include but are not limited to, mouse neuroblastoma N2A
cells, HeLa, mouse L-929 cells, 3T3 lines derived from Swiss, Balb-c or NIH mice, BHK or HaK hamster cell lines, which are available from the American Type Culture Collection, Manassas, VA). Each of these cell lines is known by and available to those skilled in the art of protein expression.
Similarly useful as host cells suitable for the present invention are bacterial cells. For example, the various strains of E. coli (e.g., HB101, (ATCC No. 33694) DH5a, DH10, and MC1061 (ATCC No.
53338) are well-known as host cells in the field of biotechnology. Various strains of B. subtilis, Pseudomonas spp., other Bacillus spp., Streptomyces spp., and the like may also be employed in this method.
Many strains of yeast cells known to those skilled in the art are also available as host cells for expression of the polypeptides of the present invention. Preferred yeast cells include, for example, saccharomyces cerivisae.
Additionally, where desired, insect cell systems may be utilized in the methods of the present invention. Such systems are described for example in Kitts et al., Biotechniques, 14:810-817 (1993), Lucklow, Curr. Opin. Biotechnol., 4:564-572 (1993) and Lucklow et al., J. Virol., 67:4566-4579 (1993).
Preferred insect cells are Sf-9 and Hi5 (Invitrogen, Carlsbad, CA).
Transformation or transfection of a nucleic acid molecule encoding an anti-IFNy antibody or antigen binding domain into a selected host cell may be accomplished by well known methods including methods such as calcium chloride, electroporation, microinjection, lipofection or the deae-dextran method.
The method selected will in part be a function of the type of host cell to be used. These methods and other suitable methods are well known to the skilled artisan, and are set forth, for example, in Sambrook et al., supra.
One may also use transgenic animals to express glycosylated selective binding agents, such as antibodies and antigen binding domain. For example, one may use a transgenic milk-producing animal (a cow or goat, for example) and obtain glycosylated binding agents in the animal milk. Alternatively, one may use plants to produce glycosylated selective binding agents.
Host cells comprising (i.e., transformed or transfected) an expression vector encoding a selective binding agent of IFNy may be cultured using standard media well known to the skilled artisan. The media will usually contain all nutrients necessary for the growth and survival of the cells. Suitable media for culturing E. co/i cells are for example, luria broth (LB) and/or terrific broth (TB). Suitable media for culturing eukaryotic cells are RPMI 1640, MEM, DMEM, all of which may be supplemented with serum and/or growth factors as required by the particular cell line being cultured. A suitable medium for insect cultures is Grace's medium supplemented with yeastolate, lactalbumin hydrolysate, and/or fetal calf serum as necessary.
Typically, an antibiotic or other compound useful for selective growth of transfected or transformed cells is added as a supplement to the media. The compound to be used will be dictated by the selectable marker element present on the plasmid with which the host cell was transformed. For example, where the selectable marker element is kanamycin resistance, the compound added to the culture medium will be kanamycin. Other compounds for selective growth include ampicillin, tetracycline and neomycin.
The amount of an anti-IFNI, antibody or antigen binding domain produced by a host cell can be evaluated using standard methods known in the art.
Such methods include, without limitation, Western blot analysis, SDS-polyacrylamide gel electrophoresis, non-denaturing gel electrophoresis, HPLC separation, immunoprecipitation, and/or activity assays.
Purification of an anti-IFNy antibody or antigen binding domain which has been secreted into the cell media can be accomplished using a variety of =
techniques including affinity, immunoaffinity or ion exchange chromatography, molecular sieve chromatography, preparative gel electrophoresis or isoelectric focusing, chromatofocusing, and high pressure liquid chromatography. For example, antibodies comprising a Fc region may be conveniently purified by affinity chromatography with Protein A, which selectively binds the fc region. Modified forms of an antibody or antigen binding domain may be prepared with affinity tags, such as hexahistidine or other small peptide such as FLAGTm(Eastman Kodak Co., New Haven, CT) or myc (Invitrogen) at either its carboxyl or amino terminus and purified by a one-step affinity column. For example, polyhistidine binds with great affinity and specificity to nickel, thus an affinity column of nickel (such as the Qiagene nickel columns) can be used for purification of polyhistidine-tagged selective binding agents; see e.g., Ausubel et al., eds., Current Protocols in Molecular Biology, section 10.11.8, John Wiley & Sons, New York (1993). In some instances, more than one purification step may be required.
Selective binding agents of the invention which are expressed in procaryotic host cells may be present in soluble form either in the periplasmic space or in the cytoplasm or in an insoluble form as part of intracellular inclusion bodies. Selective binding agents can be extracted from the host cell using any standard technique known to the skilled artisan. For example, the host cells can be lysed to release the contents of the periplasm/cytoplasm by french press, homogenization, and/or sonication followed by centrifugation.
Soluble forms of an anti-IFNy antibody or antigen binding domain present either in the cytoplasm or released from the periplasmic space may be further purified using methods known in the art, for example Fab fragments are released from the bacterial periplasmic space by osmotic shock techniques.
If an antibody or antigen binding domain has formed inclusion bodies, they can often bind to the inner and/or outer cellular membranes and thus will be found primarily in the pellet material after centrifugation.
The pellet material can then be treated at pH extremes or with chaotropic agent such as a detergent, guanidine, guanidine derivatives, urea, or urea derivatives in the presence of a reducing agent such as dithiothreitol at alkaline pH or tris carboxyethyl phosphine at acid pH to release, break apart, and solubilize the inclusion bodies. The soluble selective binding agent can then be analyzed using gel electrophoresis, immunoprecipitation or the like. If it is desired to isolate a solublized antibody or antigen binding domain, isolation may be accomplished using standard methods such as those set forth below and in Marston et al., Meth. Ehz., 182:264-275 (1990).
In some cases, an antibody or antigen binding domain may not be biologically active upon isolation.
Various methods for "refolding" or converting the polypeptide to its tertiary structure and generating disulfide linkages, can be used to restore biological activity. Such methods include exposing the solubilized polypeptide to a pH usually above 7 and in the presence of a particular concentration of a chaotrope. The selection of chaotrope is very similar to the choices used for inclusion body solubilization, but usually the chaotrope is used at a lower concentration and is not necessarily the same as chaotropes used for the solubilization. In most cases the refolding/oxidation solution will also contain a reducing agent or the reducing agent plus its oxidized form in a specific ratio to generate a particular redox potential allowing for disulfide shuffling to occur in the formation of the protein's cysteine bridge(s).
Some of the commonly used redox couples include cysteine/cystamine, glutathione (GSH)/dithiobis GSH, cupric chloride, dithiothreitol (DTT)/dithiane DTT, and 2-mercaptoethanol(bME)/dithio-b(ME). In many instances, a cosolvent may be used or may be needed to increase the efficiency of the refolding and the more common reagents used for this purpose include glycerol, polyethylene glycol of various molecular weights, arginine and the like.
Antibodies and antigen binding domains of the invention may also be prepared by chemical synthesis methods (such as solid phase peptide synthesis) using techniques known in the art such as those set forth by Merrifield et al., J. Am. Chem. Soc., 85:2149 (1963);
Houghten et al., Proc Natl Acad. Sci. USA, 82:5132 (1985), and Stewart and Young (Solid Phase Peptide Synthesis, Pierce Chemical Co., Rockford, IL (1984).
Such polypeptides may be synthesized with or without a methionine on the amino terminus. Chemically synthesized antibodies and antigen binding domains may be oxidized using methods set forth in these references to form disulfide bridges. Antibodies so prepared will retain at least one biological activity associated with a native or recombinantly produced anti-opgbp antibody or antigen binding domain.
Assays for selective binding agents of IFNy Screening methods for identifying selective binding agents which partially or completely inhibits at least one biological activity of IFNy are provided by the invention. Inhibiting the biological activity of IFNy includes, but is not limited to, inhibiting binding of IFNy to its cognate receptor, IFNy-R, inhibiting anti-proliferative activity of IFNy on A549 cells in vitro, and inhibiting activation of monocytes by IFNy in vitro and in vivo. Selective binding agents of the invention include anti-IFNy antibodies, and fragments, variants, derivatives and fusion thereof, peptides, peptidomimetic compounds or organo-mimetic compounds.Screening methods for identifying selective binding agents which can partially or completely inhibit a biological activity of IFNy can include in vitro or in vivo assays. In vitro assays include those that detect binding of IFNy to IFNy-R and may be used to screen selective binding agents of IFNy for their ability to increase or decrease the rate or extent of IFNy binding to IFNy-R. In one type of assay, an IFNy polypeptide, preferably a soluble form of IFNy such as an extracellular domain, is immobilized on a solid support (e.g., agarose or acrylic beads) and an IFNy-R
polypetpide is the added either in the presence or absence of a selective binding agent of IFNy. The extent of binding of IFNy and IFNy-R with or without a selective binding agent present is measured. Binding can be detected by for example radioactive labeling, fluorescent labeling or enzymatic reaction.
Alternatively, the binding reaction may be carried out using a surface plasmon resonance detector system such as the BIAcore assay system (Pharmacia, Piscataway, NJ). Binding reactions may be carried out according to the manufacturer's protocol.
In vitro assays such as those described above may be used advantageously to screen rapidly large numbers of selective binding agents for effects on binding of IFNy to IFNy-R. The assays may be automated to screen compounds generated in phage display, synthetic peptide and chemical synthesis libraries.
Selective binding agents increase or decrease binding of IFNy to IFNy-R may also be screened in cell culture using cells and cell lines expressing either polypeptide. Cells and cell lines may be obtained from any mammal, but preferably will be from human or other primate, canine, or rodent sources. As an example, the binding of IFNy to cells expressing IFNy-R on the surface is evaluated in the presence or absence of selective binding agents and the extent of binding may be determined by, for example, flow cytometry using a biotinylated antibody to IFNy.
In vitro activity assays may also be used to identify selective binding agents which inhibit IFNy activity. Examples of assays include A549 cell proliferation assay and THP-1 HLA-DR expression assay.

In vivo assays are also available to determine whether a selective binding agent is capable of delaying development of proteinurea and increasing survival time in NZB x NZW Fl mouse model.
For diagnostic applications, in certain embodiments, selective binding agents of IFNy, such as antibodies and antigen binding domains thereof, typically will be labeled with a detectable moiety.
The detectable moiety can be any one which is capable of producing, either directly or indirectly, a detectable signal. For example, the detectable moiety may be a radioisotope, such as 31-I, 14C, 32P, 35S, or 1251, a fluorescent or chemiluminescent compound, such as fluorescein isothiocyanate, rhodamine, or luciferin; or an enzyme, such as alkaline phosphatase, galactosidase or horseradish peroxidase; Bayer et. al., Meth. Enz., 184:138-163 (1990).
The selective binding agents of the invention may be employed in any known assay method, such as radioimmunoassays, competitive binding assays, direct and indirect sandwich assays (ELISAs), and immunoprecipitation assays (Sola, Monoclonal Antibodies: A Manual of Techniques, pp. 147-158 (CRC
Press, 1987)) for detection and quantitation of IFNy polypeptides. The antibodies will bind IFNy polypeptides with an affinity which is appropriate for the assay method being employed.
The selective binding agents of the invention also are useful for in vivo imaging, wherein for example a selective binding agent labeled with a detectable moiety is administered to an animal, preferably into the bloodstream, and the presence and location of the labeled antibody in the host is assayed. The agent may be labeled with any moiety that is detectable in an animal, whether by nuclear magnetic resonance, radiology, or other detection means known in the art.
The invention also relates to a kit comprising a selective binding agent of IFNy, such as an antibody or antigen binding domain, and other reagents useful for detecting IFNy levels in biological samples. Such reagents may include a secondary activity, a detectable label, blocking serum, positive and negative control samples, and detection reagents.
Therapeutic Uses of IFNy selective binding agents Selective binding agents of the invention may be used as therapeutics. Therapeutic selective binding agents may be IFNy agonists or antagonists and, in one embodiment, are anti-IFNy antagonist antibodies which inhibit at least one of the biological activities of an IFNy polypeptide in vitro or in vivo. For example, an antagonist of IFNy will inhibit the binding of IFNy to IFNy-R. Alternatively, an IFNy antagonist will stimulate the proliferation of human lung carcinoma in vitro as indicated by measurable ND50 (a concentration giving 50% proliferation) in a A549 cell proliferation assay such as that described in Example 1.
IFNy antagonists, such as anti-IFNy antagonist antibodies and antigen binding domains, may be used to prevent or treat auto-immune diseases and inflammatory conditions including, but not limited to the following: acute pancreatitis; ALS; Alzheimer's disease; cachexia/anorexia, including AIDS-induced cachexia; asthma and other pulmonary diseases;
atherosclerosis; chronic fatigue syndrome; Clostridium associated illnesses, including Clostridium-associated diarrhea; coronary conditions and indications, including congestive heart failure, coronary restenosis, myocardial infarction, myocardial dysfunction (e.g., related to sepsis), and coronary artery bypass graft; cancer, such as multiple myeloma and myelogenous (e.g., AML and CML) and other leukemias, as well as tumor metastasis; fever;
glomerulonephritis; graft versus host disease/transplant rejection; hemohorragic shock;
inflammatory eye disease, as may be associated with, for example, corneal transplant; ischemia, including cerebral ischemia (e.g., brain injury as a result of trauma, epilepsy, hemorrhage or stroke, each of which may lead to neurodegeneration); learning impairment;
multiple sclerosis; myopathies (e.g., muscle protein metabolism, esp. in sepsis); neurotoxicity (e.g., as induced by HIV); osteoporosis; pain, including cancer-related pain; Parkinson's disease; periodontal disease;
neurotoxicity; pre-term labor; psoriasis; reperfusion injury; septic shock; side effects from radiation therapy; temporal mandibular joint disease; sleep disturbance; uveitis; or an inflammatory condition resulting from strain, sprain, cartilage damage, trauma, orthopedic surgery, infection or other disease processes; diabetes, including juvenile onset Type 1, diabetes mellitus, and insulin resistance (e.g., as associated with obesity); endometriosis, endometritis, and related conditions; fibromyalgia or analgesia;
hyperalgesia; inflammatory bowel diseases, including Crohn's disease; lung diseases (e.g., adult respiratory distress syndrome, and pulmonary fibrosis);
neuroinflammatory diseases; ocular diseases and conditions, including ocular degeneration and uveitis;
Pityriasis rubra pilaris (PRP); prostatitis (bacterial or non-bacterial) and related conditions; psoriasis and related conditions; pulmonary fibrosis; reperfusion injury; inflammatory conditions of a joint and rheumatic diseases, including, osteoarthritis, rheumatoid arthritis, juvenile (rheumatoid) arthritis, seronegative polyarthritis, ankylosing spondylitis, Reiter's syndrome and reactive arthritis, Still's disease, psoriatic arthritis, enteropathic arthritis, polymyositis, dermatomyositis, scleroderma, systemic sclerosis, vasculitis (e.g., Kawasaki's disease), cerebral vasculitis, Lyme disease, staphylococcal-induced ("septic") arthritis, Sjogren's syndrome, rheumatic fever, polychondritis and polymyalgia rheumatica and giant cell arteritis; septic shock;
systemic lupus erythematosus (SLE) nephritis; side effects from radiation therapy; temporal mandibular joint disease; thyroiditis; tissue transplantation or an inflammatory condition resulting from strain, sprain, cartilage damage, trauma, and orthopedic surgery.
More specifically, the IFNy antagonists, such as anti-IFNy antagonist antibodies and antigen binding domains, may be used to prevent or treat arthritis (particularly rheumatoid arthritis), systemic lupus erythematosus (SLE), graft versus host disease (GvHD), multiple sclerosis and diabetes.
IFNy antagonists of the invention, including antagonist antibodies and antigen binding domains, are administered alone or in combination with other therapeutic agents IFNy antagonists, such as anti-IFNy antagonist antibodies and antigen binding domains, may be used to prevent or treat to treat various inflammatory conditions, autoimmune conditions, and other conditions leading to bone loss. Depending on the condition and the desired level of treatment, two, three, or more agents may be administered. These agents may be provided together by inclusion in the same formulation or inclusion in a treatment kit, or they may be provided separately. When administered by gene therapy, the genes encoding the protein agents may be included in the same vector, optionally under the control of the same promoter region, or in separate vectors. Particularly preferred molecules in the aforementioned classes are as follows.
= IL-1 inhibitors: IL-1ra proteins and soluble IL-1 receptors. The most preferred IL-1 inhibitor is anakinra.
= TNF-ainhibitors: soluble tumor necrosis factor receptor type I (sTNF-RI; -RI is also called the p55 receptor); soluble tumor necrosis factor receptor type II (also called the p75 receptor);
and monoclonal antibodies that bind the TNF
receptor. Most preferred is sTNF-RI as described in WO 98/24463, etanercept (Enbrel), and Avakine.
Exemplary TNF-ainhibitors are described in EP 422 339, EP 308 378, EP 393 438, EP 398 327, and EP
418 014.
= serine protease inhibitors: SLPI, ALP, MPI, HUSI-I, BMI, and CUSI. These inhibitors also may be viewed as exemplary LPS modulators, as SLPI has been shown to inhibit LPS responses. Jin et al.
(1997), Cell 88(3): 417-26.
Pharmaceutical Compositions PhaLmaceutical compositions of IFNy selective binding agents are within the scope of the present .
invention. Such compositions comprise a therapeutically or prophylactically effective amount of an IFNy selective binding agent such as an antibody, or a fragment, variant, derivative or fusion thereof, in admixture with a pharmaceutically acceptable agent. In a preferred embodiment, pharmaceutical compositions comprise anti-IFNrantagonist antibodies which inhibit partially or completely at least one biological activity of IFNy in admixture with a pharmaceutically acceptable agent. Typically, the antibodies will be sufficiently purified for administration to an animal.
Pharmaceutically acceptable agents for use in the compositions of the invention include carriers, excipients, diluents, antioxidants, preservatives, coloring, flavoring and diluting agents, emulsifying agents, suspending agents, solvents, fillers, bulking agents, buffers, delivery vehicles, tonicity agents, cosolvents, wetting agents, complexing agents, buffering agents, antimicrobials and surfactants, as are well known in the art.
Neutral buffered saline or saline mixed with serum albumin are exemplary appropriate carriers. Also included in the compositions are antioxidants such as ascorbic acid; low molecular weight polypeptides;
proteins, such as serum albumin, gelatin, or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, arginine or lysine;
monosaccharides, disaccharides, and other carbohydrates including glucose, mannose, or dextrins; chelating agents such as EDTA; sugar alcohols such as mannitol or sorbitol; salt-forming counterions such as sodium;
and/or nonionic surfactants such as Tween, pluronics or polyethylene glycol. Also by way of example, suitable tonicity enhancing agents include alkali metal halides (preferably sodium or potassium chloride), mannitol, sorbitol and the like. Suitable preservatives include, but are not limited to, benzalkonium chloride, thimerosal, phenethyl alcohol, methylparaben, propylparaben, chlorhexidine, sorbic acid and the like.

Hydrogen peroxide may also be used as preservative.
Suitable cosolvents are for example glycerin, propylene glycol, and polyethylene glycol. Suitable complexing agents are for example caffeine, polyvinylpyrrolidone, beta-cyclodextrin or hydroxy-propyl-beta-cyclodextrin. Suitable surfactants or wetting agents include sorbitan esters, polysorbates such as polysorbate 80, tromethamine, lecithin, cholesterol, tyloxapal and the like. The buffers can be conventional buffers such as acetate, borate, citrate, phosphate, bicarbonate, or Tris-HC1. Acetate buffer may be around pH 4.0-5.5 and Tris buffer may be around pH 7.0-8.5. Additional pharmaceutical agents are set forth in Remington's Phaimaceutical Sciences, 18th Edition, A.R. Gennaro, ed., Mack Publishing Company 1990.
The compositions may be in liquid form or in a lyophilized or freeze-dried form. Lypophilized forms may include excipients such as sucrose.
The compositions of the invention are suitable for parenteral administration. In preferred embodiments, the compositions are suitable for injection or infusion into an animal by any route available to the skilled worker, such as subcutaneous, intravenous, intramuscular, intraperitoneal, intracerebral (intraparenchymal), intracerebroventricular, intramuscular, intraocular, intraarterial, or intralesional routes. A parenteral formulation will typically be a sterile, pyrogen-free, isotonic aqueous solution, optionally containing pharmaceutically acceptable preservatives.
The optimal pharmaceutical formulation may be readily determined by one skilled in the art depending upon the intended route of administration, delivery format and desired dosage.

Other formulations are also contemplated by the invention. The pharmaceutical compositions also may include particulate preparations of polymeric compounds such as polylactic acid, polyglycolic acid, etc. or the introduction of an IFNy selective binding agent (such as an antibody) into liposomes. Hyaluronic acid may also be used, and this may have the effect of promoting sustained duration in the circulation.
Pharmaceutical compositions also include the formulation of IFNy selective binding agents (such as antibodies) with an agent, such as injectable microspheres, bio-erodible particles or beads, or liposomes, that provides for the controlled or sustained release of a selective binding agent which may then be delivered as a depot injection. Other suitable means for delivery include implantable delivery devices.
A pharmaceutical composition comprising and IFNy selective binding agent (such as an antibody) may be formulated as a dry powder for inhalation. Such inhalation solutions may also be formulated in a liquefied propellant for aerosol delivery. In yet another formulation, solutions may be nebulized.
It is also contemplated that certain formulations containing IFNy selective binding agents may be administered orally. Formulations administered in this fashion may be formulated with or without those carriers customarily used in the compounding of solid dosage forms such as tablets and capsules. For example, a capsule may be designed to release the active portion of the formulation at the point in the gastrointestinal tract when bioavailability is maximized and pre-systemic degradation is minimized.
Additional agents may be included to facilitate absorption of a selective binding agent. Diluents, flavorings, low melting point waxes, vegetable oils, lubricants, suspending agents, tablet disintegrating agents, and binders may also be employed.
Another preparation may involve an effective quantity of an IFNy selective binding agent in a mixture with non-toxic excipients which are suitable for the manufacture of tablets. By dissolving the tablets in sterile water, or another appropriate vehicle, solutions can be prepared in unit dose faun.
Suitable excipients include, but are not limited to, inert diluents, such as calcium carbonate, sodium carbonate or bicarbonate, lactose, or calcium phosphate; or binding agents, such as starch, gelatin, or acacia; or lubricating agents such as magnesium stearate, stearic acid, or talc.
Additional formulations will be evident to those skilled in the art, including formulations involving IFNy selective binding agents in combination with one or more other therapeutic agents. Techniques for formulating a variety of other sustained- or controlled-delivery means, such as liposome carriers, bio-erodible microparticles or porous beads and depot injections, are also known to those skilled in the art.
See, for example, the Supersaxo et al. description of controlled release porous polymeric microparticles for the delivery of pharmaceutical compositions (See WO
93/15722 (PCT/US93/00829).
Regardless of the manner of administration, the specific dose may be calculated according to body weight, body surface area or organ size. Further refinement of the calculations necessary to determine the appropriate dosage for treatment involving each of the above mentioned formulations is routinely made by those of ordinary skill in the art and is within the ambit of tasks routinely performed by them.
Appropriate dosages may be ascertained through use of appropriate dose-response data.
One may further administer the present pharmaceutical compositions by pulmonary administration, see, e.g., PCT W094/20069, which discloses pulmonary delivery of chemically modified proteins. For pulmonary delivery, the particle size should be suitable for delivery to the distal lung. For example, the particle size may be from lAm to 5Am, however, larger particles may be used, for example, if each particle is fairly porous.
Alternatively or additionally, the compositions may be administered locally via implantation into the affected area of a membrane, sponge, or other appropriate material on to which an IFNy selective binding agent has been absorbed or encapsulated. Where an implantation device is used, the device may be implanted into any suitable tissue or organ, and delivery of an IFNy selective binding agent may be directly through the device via bolus, or via continuous administration, or via catheter using continuous infusion.
Pharmaceutical compositions of the invention may also be administered in a sustained release formulation or preparation. Suitable examples of sustained-release preparations include semipermeable polymer matrices in the form of shaped articles, e.g.
films, or microcapsules. Sustained release matrices include polyesters, hydrogels, polylactides (See e.g., U.S. Patent No. 3,773,919, EP 58,481), copolymers of L-glutamic acid and gamma ethyl-L-glutamate (Sidman et al, Biopolymers, 22: 547-556 (1983), poly (2-hydroxyethyl-methacrylate) (Langer et al., J. Biamed.

Mater. Res., 15: 167-277 (1981) and Langer, Chem.
Tech., 12: 98-105 (1982), ethylene vinyl acetate, or poly-D(-)-3-hydroxybutyric acid. Sustained-release compositions also may include liposomes, which can be prepared by any of several methods known in the art.
See e.g., Eppstein et al., Proc. Natl. Acad. Sci. USA, 82: 3688-3692 (1985); EP 36,676; EP 88,046; and EP
143,949.
It may be desirable in some instances to use a pharmaceutical composition comprising an IFNy selective binding agent compositions in an ex vivo manner. Here, cells, tissues, or organs that have been removed from the patient are exposed to pharmaceutical compositons comprising IFNy selective binding agents after which the cells, tissues and/or organs are subsequently implanted back into the patient.
In other cases, a composition comprising an IFNy selective binding agent may be delivered through implanting into patients certain cells that have been genetically engineered, using methods such as those described herein, to express and secrete the polypeptides, selective binding agents, fragments, variants, or derivatives. Such cells may be animal or human cells, and may be derived from the patient's own tissue or from another source, either human or non-human. Optionally, the cells may be immortalized.
However, in order to decrease the chance of an immunological response, it is preferred that the cells be encapsulated to avoid infiltration of surrounding tissues. The encapsulation materials are typically biocompatible, semi-permeable polymeric enclosures or membranes that allow release of the protein product(s) but prevent destruction of the cells by the patient's immune system or by other detrimental factors from the surrounding tissues.

Methods used for membrane encapsulation of cells are familiar to the skilled artisan, and preparation of encapsulated cells and their implantation in patients may be accomplished without undue experimentation. See, e.g., U.S. Patent Nos.
4,892,538, 5,011,472, and 5,106,627. A system for encapsulating living cells is described in PCT WO
91/10425 (Aebischer et al.). Techniques for formulating a variety of other sustained or controlled delivery means, such as liposome carriers, bio-erodible particles or beads, are also known to those in the art, and are described. The cells, with or without encapsulation, may be implanted into suitable body tissues or organs of the patient.
A therapeutically or prophylactically effective amount of a pharmaceutical composition comprising an IFNy selective binding agent (such as an anti-IFNy antibody, or fragment, variant, derivative, and fusion thereof) will depend, for example, upon the therapeutic objectives such as the indication for which the composition is being used, the route of administration, and the condition of the subject. IFNy antagonist antibodies or antigen binding domains of the invention are administered in a therapeutically or prophylactically effective amount to prevent and/or treat an auto-immune and/or inflammatory condition.
The following examples are offered to more fully illustrate the invention but are not construed as limiting the scope thereof.
Example 1 Reagents and Assays The screening targets used in these studies were hIFNy prepared from: 1) expression of a cDNA
encoding hIFNy in E. coii as described in EP 0423845, or PCT Publication WO 83/04053; or, 2) expression of a cDNA encoding hIFNy in a CHO host cell as follows: PCR
(standard conditions) was used to amplify the full-length sequence encoding the human IFNy using human spleen marathon ready cDNA (Clontech) as a template.
The sequence was subcloned into an expression plasmid and DNA transformed into DH1OB cells (Gibco Life Sciences), DNA prepared, and transfected into CHO cells by the calcium phosphate method (Speciality Media, Inc.). A high-expressing cell line clone was used to generate serum-free conditioned media.
CHO cell conditioned media containing hINFy was concentrated, dialyzed, and then purified through several chromatography steps. The lst step was Q-HP
(Pharmacia) chromatography using a standard NaC1 gradient to separate highly glycosylated vs.
unglycosylated hIFNy forms. The Q-HP pool was further purified through a wheat germ agglutinin chromatography (EY Laboratories). The purified material was greater than 95% pure judged by both Coomassie-blue and silver-stained SDS-PAGE. The material was of low endotoxin level as assayed by the gel-clot method (Limulus Amebocyte Lysate). The identity of hINFy was confirmed by western blot, using goat anti-hIFNy neutralizing antibody from R & D Systems (catalog number AF-285-NA, lot number ZW019011). The final protein concentration was determined using the extinction coefficient method (0.66). Two lots of material were generated respectively. The yield was 40mg/l. Final materials were formulated in PBS.

Expression of human IFNyR1-Fc protein in CHO cells The human IFNyR1-Fc protein used for elution of phage antibodies from target in these studies were prepared as follows: PCR (standard conditions) was used to amplify the full-length sequence encoding the human IFNyR1 using human lymphoid marathon ready cDNA
(purchased from Clontech) as a template. PCR (standard conditions) was used to amplify the sequence encoding the Fc portion of human IgG1. Overlap PCR was used to generate a sequence encoding the IFNyR1-Fc fusion construct (Amino acids 1- Ser2" of the IFNyR1) and the sequence was subcloned into an expression plasmid. DNA
was transformed into DH1OB cells (Gibco Life Sciences), DNA prepared, and transfected into CHO cells by the calcium phosphate method (Speciality Media, Inc). A
high-expressing cell line clone was used to generate serum-free conditioned media.
CHO cell conditioned media containing hINFyR1-Fc was concentrated and purified through standard Protein-G Fast-Flow column (Pharmacia). Final concentration was determined by A,. using 1.44 as the extinction coefficient. The identity of the purified sample was confirmed through N-terminal sequencing analysis. The material was formulated in PBS.
Antibodies Monoclonal anti-hIFNy antibody, clone 2578.111, was purchased from R&D Systems (catalog number MAB285, lot number KW07). Monoclonal anti-hIFNy antibody, clone MMHG-1, was purchased from Biosource (catalog number AHC4834, lot number 10803-015).
Recombinant human IFNy Receptorl (rhIFNy R1) was purchased from R&D Systems (catalog number 673-IR).
The calculated molecular weight of the rhIFNy R1 is 25,000 daltons. As a result of glycosylation, the recombinant protein migrates as a 40-50 kDa protein on SDS-PAGE.
A549 cell proliferation assay The A549 cell proliferation assay used to evaluate antibody neutralization of IFNy is a 96 well assay and is generally described as follows: on day 1, 1) dilute Ab serially 1:2 from highest concentration in Assay Media (F12K, 5% FBS, lx Pen/strep L-Glutamine). Do a total of 10 dilutions at 4X the concentration desired in Assay. For duplicates, at least 200 1 final is needed for each dilution; 2) dilute IFNy to appropriate concentration for spike, based on 90% of the effective dose in a dose response curve. Make IFNy spike 4X the concentration desired in assay; 3) combine 150 1 of each 4XAb dilution with 150 1 4X IFNy spike in titertek tubes. Mix by pipetting. Cover and incubate 1 hour at room temperature. (Note: Concentration of Ab and IFNy now at 2X assay concentration); 4)(Optional) while Ab and IFNy incubate, dilute IFNy for titration curve. Do 12 1:3 dilutions starting at 4000 ng/ml. Since 300 1 is needed for triplicates in assay, volume needed at end of dilution should be at least 400 1. Store at 4 C
until needed; 5) before incubation is completed, trypsinize A549 cells in 5m1 trypsin. Add 20m1 Assay Media to flask and transfer to 50m1 conical and centrifuge at IA to 3/4 speed in IEC, RT; 6) aspirate cells. Resuspend in 7.5 ml Assay Media. Count 1:1 in trypan Blue; 7) dilute Cells in assay media to 2.5 X
104cells/ml. Seed 0.1 ml into 96 well falcon for each sample for 2.5 X 10'cells/well; 8) at one hour, add 100111 Ab/IFNy mix to each of two wells for duplicates;

and 9) incubate for 5 days at 37 C, 5% CO2 and high humidity. On day 5: 1) add 20 1 Alamar Blue per well.
Incubate for 3-4 hours at 37 C, 5% CO2 and high humidity; 2) turn on FL500 fluorescent plate reader.
Remove lids from plates and shake for 10 minutes, without lid; and 3) read on FL500. Settings: Shake 3 seconds at medium, excitation at 530/25, emission at 590/35, sensitivity of 34.
Example 2 Screening of a Human Fab library Screening procedure General procedures for construction and screening human Fab libraries were described in de Haard et al. (Advanced Drug Delivery Reviews, 31:5-31 (1998); J. Biol. Chem., 274:18218-18230 (1999)). The library was screened for Fab fragments which bind to hIFNy by the following procedures.
Nunc immunotube was coated with 4 ml of hIFNI/
at 0.39 g/ml in 0.1 M Na carbonate, pH 9.6 at room temperature on Nutatorrmfor 2 hrs. After thawing, glycerol (15%) was removed from an aliquot of Target Quest, NV (Amsterdam, Netherlands) frozen phage library stock (4x 1012pfuin 750 1 per tube) by adding 1/5 vol.
(150 1) of PEG solution (20% polyetheylene glycol 8000, 2.5 M NaC1, autoclaved) and leaving the tube on ice for 1 hr to precipitate the phage. The precipitated phage particles were pelleted at 4000 rpm for 15 min at 4 C, then resuspended into 500 1 PBS, pH
7.4. IFNy-coated immunotube was washed 3Xs with 4 ml PBS and blocked with 4 ml 2% MPBS at RT for 1 hr on Nutator. At the same time, 500 Al 4% MPBS was added to the phage suspension and incubated for 30 min-1 hr at room temperature to allow pre-blocking of the phage particles. The blocked immunotube was washed with 2x TM
PBST(0.1% Tween20 in PBS) and 2x with PBS. The pre-blocked phage mixture was added to the washed immunotube containing 3 ml of 2% MPBS. After 30 minutes of incubation on a rotator followed by 1.5 hr of standing incubation at room temperature, the phage mixture was discarded. The tube was washed first 20x with PBST, then 20x with PBS. The bound phage particles were eluted by incubation with 1 ml of specific elution reagent (hIFN/, GPNA, RDMA, BSMA, or rhIFNy R1, respectively) at 1 M in 0.4% MPBS, pH 7.4 for 90 man on a rotator. The eluted phage particles were transferred to sterile 50 ml conical polypropylene tube and stored on ice. About 20 1 of each phage elution was set aside for titering. For amplification, the remaining eluted phage particles were added to a 50 ml conical tube containing 5 ml of TG1 culture (0D590 about 0.5) and 4 ml 2x YT. The IFNection mixture was incubated at 37 C without shaking for 30 min, then spun at 3500 rpm for 20 min. The cell pellet was suspended into 1500 1 2xYT-AG broth and plated 300 1/plate on five SOBCG plates. The plates were incubated at 30 C overnight. After 20 hours of incubation, the cells were recovered with cell scraper from the plates, to which 4 ml per plate of 2x YT-AG
were added. The step was repeated three times. A
small portion of the recovered cells was used for phage rescue (see below). The remaining cell suspension .was =
spun at 3500 rpm for 20 man. The cell pellet was suspended into %volume of the pellet size of 50%
glycerol to make glycerol stocks and stored at -80 C.
Phage rescue from amplified cell suspension was performed as follows. About 0.5 ml of recovered plated-amplified cell suspension was used to inoculate 50 ml of 2x YT-AG to 0D590 about 0.3. The culture was incubated at 37 C on a shaker to 0D590 0.5. 10 ml of the culture was IFNected with 1 ml of M13K07 helper phage (GIBCO BRL, catalog # 18311-019, 1.1 x 1011 pfu/ml) at M.O.I. 20. and incubate in the incubator at 37 C for 30 min. The IFNected cells were spun down at 4000 rpm for 20 min. The cell pellet was re-suspended into 50 ml of 2xYT-AK, transferred to a 250-ml flask and incubated at 30 C with shaking at 270 rpm for 20 hours. The over-night culture was spun at 4000 rpm for min to removal cell debris. The supernatant was centrifuge again to ensure the removal of cell debris.
About 1/5 volume of PEG solution (20% PEG 8000, 2.5 M
15 NaCl) was added to the supernatant to precipitate the phage particles. The mixture was incubated on ice for at least 1 hour, then centrifuged at 4000 rpm for 20 min to collect the precipitated phage particles. The phage pellet was re-suspended into 1 ml of PBS and 20 transferred to a microfuge tube. The phage suspension was left on ice for 1 hour to allow complete suspension of phage particles, then spun at 14,000 rpm for 2 min to remove the residual cell debris. Phage precipitation step was repeated. The final phage pellet was suspended into 1.1 ml of PBS and left on ice for an extended period to ensure complete suspension of phage particles. The phage suspension was centrifuged at 14,000 rpm for 2 min to remove residual cell debris.
500 1 of rescued phage suspension was used to make a glycerol stock by addition of 250 1 of 50% glycerol.
100 1 of rescued phage suspension was reserved for phage pool ELISA (see below). The remaining 500 1 of the rescued phage was used for next round of panning.

Phage pool ELISA
Phage pool ELISA was performed as follows:
E. coil expressed hIFNy was plated, 100 l/well, at 0.39 g/ml in 0.1 M Na carbonate, pH 9.6 in Nunc MaxiSorb Immuno plate at room temperature with gentle rocking for 2 hrs. The coated plate was washed 3 times with PBS, then blocked with 300 l/well of 2% MPBS at room temperature on the rocker for one hour. For negative control, another Nunc Immuno plate which has not been coated with the antigen was also blocked with 2% MPBS. Meanwhile, 120 1 of each rescued phage pool with was pre-blocked with 120 1 of 0.8%MTBS in a 96-well Costar 3790 plate and left at room temperature until ready to use. Both blocked plates were washed 5 times with 0.1%TBST (TBS: 10 mM Tris-HC1, pH 7.5, 1 mM
EDTA, 150 mM NaCl; Tween-20. 0.1%). Pre-blocked phage dilution was distributed (100 l/well) to both antigen-coated plate and the negative control plate, and incubated at room temperature on rocker for one hour.
After the plates were washed as described, 100 1/well of 1:1000 fold diluted HRP/Anti-M13 monoclonal Conjugate (Amersham Pharmacia Biotech, catalog number 27-9421-01) in 0.4% MTBS was distributed, and incubated at room temperature on rocker for one hour. The plates were washed as described. After 100 l/well of the substrate l_StepTM ABTS (Pierce, catalog number 37615) was added, the plates were incubated for one hour.
OD,, was measured for signal detection. ELISA positive phage pools were used as the source of individual clones for further analysis.
Example 3 Identification of IFNy binding Fab phage clones DNA Fingerprinting Polymerase chain reaction (PCR) was performed in a 96-well Thermowell plate in order to identify full-length clones containing both heavy chain and light chain from ELISA positive phage pools.
Typically, each well contains 25 1 of PCR reaction mix (2.5 1 10X PCR buffer, 21.625 1 water, 0.25 1 dNTPs at 25 mM, 0.25 1 primer 870-02 (shown below) at 10 pmol/ 1, 0.25 1 primer 2182-83 (shown below) at 10 pmol/ 1, 0.125 1 Tag polymerase at 5 units/ 1).
870-02 5'-CCG ACT TTG CAC CTA GTT (SEQ ID NO:109) 2182-83 5'-TTT GTC GTC TTT CCA GAC GTT AGT
(SEQ ID NO:110) Individual colonies were picked and resuspended first into a well in the PCR plate, then resuspended into the corresponding well in a 96-deep well block filled with 300 l/well of 2xYT-AG broth (2xYT broth: 10 g yeast extract, 16 g bacto-tryptone, 5 g NaCl per liter of water containing 100 g/ml ampicillin and 2% glucose).
The PCR reaction conditions were one denature cycle of 5 min at 94 C, 40 cycles of 45 sec at 94 C, 45 sec at 55 C, 1.5 min at 72 C, followed by one extension cycle at 72 C for 10 min. After completion of PCR
reaction, 3 l/well of PCR reaction mixture were run on a 1% extra long 4x(24+2) TAE gels containing 0.5u1/m1 ethidium bromide (Embi Tec, catalog # GE-3820) at 120 volts for one hour. By comparison to the 1 kb plus DNA
ladder (Gibco BRL, catalog # 10787-018), clones with inserts greater than 1.6 kb were identified as full-length clones.

Identification of unique full-length clones was performed as follows: BstNI digestion was performed on PCR amplified inserts of the identified full-length clones. To 16 1 of PCR reaction mixture per sample in a 96-well Thermowell plate, 14 1 of BstNI digestion master solution containing 3 1 10x Buffer 2 (NEBL), 0.3 1 BSA at 10 mg/ml, 10 1 water and 0.7 1 BstNI (NEBL) was added. The plates were incubated at 60 C for 3 hours. Digested samples, 13 1 each, were run on 4% extra long 2x(24+2) TAE gels containing 0.5u1/m1 ethidium bromide (Embi Tec, catalog # GE-3817) at 100 volts for 3 hours. Unique clones were identified based on the difference in BstNI
fragment patterns.
Clonal phage ELISA
Fab phages of identified unique full-length clones were rescued in the 96-well format. In 96-well 2-ml deep-well block, 480 l/well 2xYTAG broth was inoculated with 20 1 of overnight cultures of the selected unique full-length clones, then incubated at 37 C, 300 rpm for 3 hours. To each well, 100 1 of 1:10 diluted M13K07 helper phage dilution were added to IFNect the cells. The block was incubated at 37 C
without shaking for 30 minutes, then shaken gently for another 30 minutes at 150 rpm. The block was centrifuged at 3600 rpm for 20 minutes to pellet the IFNected cells. The cell pellet in each well was suspended into 480 1 of 2xYTAK (2xYT broth containing 100 g/ml ampicillin and 40 g/ml kanamycin), then incubated at 30 C overnight for about 20 hours. The cell debris was separated by centrifugation at 3600 rpm for 20 minutes. The rescued phage supernatant was carefully transfer into another sterile 96-well block.
The rescued phages were used to perform clonal phage ELISA exactly the same as described in Example 2, Phage pool ELISA. Clones that give 0.2 net OD.,were considered as IFNy-binding candidates.
Large scale phaqe rescue ELISA
Specific IFNy-binding of the identified unique Fab phage clones was confirmed by demonstration of concentration-dependent Fab phage binding to IFNy in ELISA. Fab phages were obtained by large scale rescue.
Large-scale rescue of individual clones was performed as follows: Fab phages of identified unique IFNy-binding clones were rescued in large scale. In a 250-ml sterile flask, 50 ml of 2xYT-AG broth was inoculated with 200 1 of overnight culture of the selected IFNy-binding clone, and incubated at 37 C, 2700 rpm until the 0D590 of the culture reacheed 0.5.
Five ml of M13K07 helper phage (GIBCO BRL, catalog #
18311-019, 1.1 x 1011 pfu/ml) were added to infect the cells at M.O.I of 20. The cell/helper phage mixture was incubated at 37 C without shaking for 30 minutes, then centrifuged at 4000 rpm for 20 minutes to pellet the infected cells. The cell pellet was suspended in 50 ml of 2xYTAK broth (2xYT broth containing 100 g/ml ampicillin and 40 g/ml kanamycin), then incubated at C with shaking at 270 rpm overnight for about 20 hours. The overnight culture was centrifuged at 4000 30 rpm for 20 minutes to remove the cell debris. The supernatant was centrifuged again to ensure the removal of cell debris. To the supernatant, 10 ml (1/5 vol.) of PEG solution (20% PEG 8000, 2.5 M NaC1) was added to precipitate the phage particles. The mixture was incubated on ice for at least 1 hour, and centrifuged at 4000 rpm for 20 min to collect the precipitated phage particles. The phage pellet was re-suspended in 1 ml of PBS and transferred to a microfuge tube. The phage suspension was left on ice for 1 hour to allow complete suspension of phage particles, then spun at 14,000 rpm for 2 min to remove the residual cell debris. Phage precipitation step was repeated. The final phage pellet was suspended into 1 ml of PBS and left on ice for an extended period to ensure complete suspension of phage particles. The phage suspension was centrifuged at 14,000 rpm for 2 min to remove residual cell debris. The final phage suspension was stored at 4 C. Phage ELISA was performed as described in Example 2, phage pool ELISA. At least six different concentrations of large-scale rescued phages, typically from 1x109 pfu/well to 1x1011pfu/well, were added to the corresponding wells.
A total of eleven Fab clones were identified.
Fab clones "IFN-A", "57E", and "57D" were identified from phage pool with E. coli hIFN-y elution. Fab clones "GP-A" and "58C" were identified from phage pool with GPNA elution. Fab clones "RD-A2", "RD-B" and "59-A2" were identified from phage pool with RDMA elution.
Fab clones "BS-A" and "BS-B" were identified from phage pool with BSMA elution. Fab clone "67C" was identified from phage pool with hIFN-y R1 elution. Concentration dependent clonal phage ELISA of nine unique clones was performed on large-scale rescued phage preparations and illustrated in Figure 1 and Figure 2. These Fab phages can be grouped into three groups based on their ELISA
profiles. Group A includes Fab clones "GP-A" and "BS-B". These two Fab phages are strong binders, with ELISA signals reaching saturation at 5E9 pfu/well.

Group B includes Fab clones "BS-A", "RD-A2", "INF-A"
and "57E". These are strong to moderate binders that show good concentration-dependent binding curves.
Group C includes Fab clones "57D", "58C", "RD-B" and "67C". These are weak yet specific and concentration-dependent binders of IFNy.
Sequence Analysis of Fab clones Confirmation of unique IFNy binding Fab phage clones was performed as follows: Plasmid DNAs of representatives of each unique Fab BstNI digestion pattern were prepared using QIAfilterTM Plasmid midi kit (Qiagen, catalog # 12245) and sent for sequencing.
Sequences of all Fab BstNI patterns confirmed their uniqueness and revealed their individual heavy chain and light chain sequence (see Figures 3-24).
The DNA and predicted amino acid sequences for the heavy chains of Fabs "BS-A", "BS-B", "RD-B1", "RD-A2", "58C", "GP-A", "57D", "57E", "IFN-A", "67C"
and "59-A2" (SEQ ID Nos:65-86, respectively) were shown in Figures 3-13, respectively. The DNA and predicted amino acid sequences for the light chains of Fabs "BS-A", "BS-B", "RD-B1", "RD-A2", "58C", "GP-A", "57D", "57E", "IFN-A", "67C", and "59-A2" (SEQ ID Nos:87-108, respectively) were shown in Figures 14-24, respectively. The amino acid sequences of the heavy chains and the light chains of all eleven Fabs were compared, as shown in Figure 31. GCG's "BestFit"
program was used to obtain percentage of identity and similarity between each pair of Fabs. The closest matches in the heavy chains are in "BS-A", "RD-A2"
and"IFN-A". The heavy chain sequences of "BS-A"
and"RD-A2" have identical framework and CDR1 and CDR2.
They differ only in CDR3, and have 92.6 % identity and 93.4 % similarity. With the exception of the 1st amino acid, the heavy chain sequences of "BS-A" and "IFN-A"
have identical framework and CDR1 and CDR2 and different CDR3. They share 93.5 % identity and 95.1 %
similarity. The same is true for the heavy chain sequences of "IFN-A" and "RD-A2", with identity of 90.5 % and similarity of 92.1 %. The Amino acid sequence of heavy chain of Fab "57E" shows 88.1 % identity and 89.0 % similarity to the heavy chain sequence of "BS-A", 88.8 % identity and 90.0 % similarity to the heavy chain sequence of "RD-A2", and 89.0 identity and 90.7 %
similarity to the heavy chain sequence of IFN-A. The Amino acid sequence of heavy chain of Fab "BS-B" shows 88.6 % identity and 90.4 % similarity to the heavy chain sequence of "57D", 81.7 % identity and 83.3 %
similarity to the heavy chain sequence of "GP-A", and 83.9% identity and 84.7 % similarity to the heavy chain sequence of 58C. The closet matches in the light chains are between "59-A2" and "BS-A" with 90.8 % identity and 91.7 % similarity, between "BS-A" and "BS-B" with 89.0 % identity and 90.9 % similarity, between "57E" and "BS-A" with 88.2 % identity and 90.9 % similarity, between "57E" and "BS-B" with 89.7 % identity and 91.5%
similarity, and between "59-A2" and"BS-B" with 88.2 %
identity and 88.2 % similarity. Only three pairs of Fabs, "59- A2"/"BS-B", "57E"/"BS-A" and "IFN-A"/"RD-A2", are closely matched in both heavy chain and light chain.
A comparison of amino acid sequences of complementary determining regions (CDRs) is shown in Figure 25. The heavy-chain CDR35 of the eleven anti-IFNy Fabs share little similarities. The Fabs can be grouped according to the similarities of either the heavy chain CDRs or the light chain CDRs, as shown in Figure 32. Clones "BS-A", "IFN-A" and "RD-A2" have identical heavy chain CDR1 and CDR2. However, beside the same last three residues (FDY), their heavy chain CDR3s are very different. Interestingly, IFN-A and RD-A2 also share closely matched light chain CDR1 (11/16 identical residues), CDR2 (5/7 identical resudes), and CDR3 (8/9 identical residues). Clones "BS-B", "59-A2", "GP-A", and "57D" have similar heavy chain CDR1 and CDR2. Clones "BS-B" and "59-A2" have identical heavy chain CDR1 and CDR2, yet very different heavy chain CDR3. All three light chain CDRs of clones "59-A2", "BS-A", "BS-B" and "57E" are very similar.
Example 4 Expression and Purification of Soluble Fabs E. coli strain HB2151 (Pharmacia) was transformed with plasmid DNA of a unique binder.
Overnight cultures of the transformed HB2151 were grown in 2xYT-AG broth at 30 C. 750 ml of 2xYT containing 100 g/ml amplicillin and 0.1% glucose were inoculated with 7.5 ml of overnight culture and incubated at 37 C
with shaking (270 rpm) for about 2 hours. When 0D590 reached 0.8-1.0, IPTG was added to 1 mM for induction.
The culture was continued to grow at 30 C for 4 hours while shaking. The culture was centrifuged at 4 000 rpm for 20 min and the supernatant was discarded.
Periplasmic release of Fab was achieved using Osmotic shock approach. Cells were suspended in 8 ml of ice cold TES (0.2 M Tris, 0.5 mM EDTA, 17.1% sucrose, pH
8.0) and incubated on ice for 5-10 min with occasional gentle shaking. The empty tube was rinsed with 8.8 ml TES/H20 (1:3), which was pooled to the cell suspension.
The cell suspension was incubated on ice for another 20 min, and centrifuged at 4,000 rpm for 15 min. The supernatant was carefully transferred into another tube and centrifuged again at 8000 rpm for 20 min. The resulted supernatant was the TES-released periplasmic fraction. The cell pellet was resuspended in 10 ml TES / 15 mM MgSO4, incubated on ice for 15 min, then centrifuged twice as described above. The final supernatant was the Mg-released periplasmic fraction and was pooled together with the TES-released periplasmic fraction.
BSA was added as a carrier and stabilizer to the periplasmic fraction to a final concentration of 1 mg/ml. The periplasmic fraction was dialyzed with one change against 2 L of sonification buffer (20mM Tris-HC1 / 0.1M NaC1, pH 8.5) plus protease inhibitors at 4 C. The periplasmic fraction was added to 1/10th volume of pre-equilibrated TALON resin (Clontech) and incubated at 4 C with gentle rocking for 1 hour. The resin mixture was centrifuged at 1300 rpm for 3 min, and the supernatant was removed as much as possible.
The resin was wash with 10 volumes of sonification buffer, then centrifuged at 1300 rpm for 3 min. The supernatant was discarded. The washed resin was suspended into one bed volume of sonification buffer and packed into a column, which was washed with three bed volumes of sonification buffer. The Fab was eluted with 2 bed volumns of 200mM imidazole. Purified Fab was dialyzed into PBS, pH 7.4.
Example 5 Cloning and Expression of full-length human IFNy antibodies FAb clones were converted to full-length antibodies by the following procedures.
Construction of pDSRa19:hIgG1 CH

The plasmid pDSRa19:anti human OPGL IgG1 was digested with Hind= and BsmBI to remove the coding region for anti-human OPGL variable region. The linear plasmid pDSRa19:hIgG1 CH containing the 1.0 kbp human IgG1 constant region domain (C.1, hinge, C.2 and C.3 domains) was gel isolated and used to accept FAb derived anti IFN-gamma variable regions.
Construction of pDSRa19:anti-IFN gamma BS-A heavy chain The anti-IFN-gamma FAb heavy chain cDNAs were cloned into pDSRa19:hIgG1 CH to convert the FAbs into full length IgGs. The construction of a plasmid encoding "BS-A" heavy chain is described here. The other FAb heavy chains were cloned using similar procedures. To generate the FAb with a signal sequence, a three-step PCR was performed. First, primers 2485-51 (shown below) and 2465-68 (shown below) were used with the FAb cDNA template. Conditions were:
94 C for 1 min, (94 C for 20 sec., 48 C for 30 sec., 74 C
for 30 sec.) for 4 cycles, (94 C for 20 sec.,66 C for 30 sec., 74 C for 30 sec.) for 25 cycles and 74 C for 5 min. with Pfu polymerase and the appropriate buffer and nucleotides. The PCR product was then amplified with primers 2148-98 (shown below) and 2465-68 (shown below) followed by amplification with primers 2489-36 (shown below) and 2465-68 (shown below). The final PCR
product was Qiagen purified, cut with HindIII and BsmBI, and Qiagen purified. This fragment containing the FAb with a 5' Kozak (translational initiation) site and the following signal sequence for mammalian expression:
MDMRVPAQLLGLLLLWLRGARC (SEQ ID NO: 111), was ligated into pDSRa19:hIgG1 CH.

2489-36 5'-CAG CAG AAG OTT CTA GAC CAC CAT GGA CAT
GAG GGT CCC CGC TCA GCT CCT GGG
(SEQ ID NO:112) 2148-98 5'-CCG CTC AGC TCC TGG GGC TCC TGC TAT TGT
GGT TGA GAG GTG CCA GAT (SEQ ID NO:113) 2485-51 5'-G TGG TTG AGA GGT GCC AGA TGT CAG GTG CAG
CTG CAG GAG-3' (SEQ ID NO:114) 2465-68 5'-GT GGA GGC ACT AGA GAO GGT GAO CAG GGT 3' (SEQ ID NO:115) Construction of pDSRa19: anti-IFN gamma BS-A heavy chain The FAb light chain cDNAs were cloned into pDSRa19 to convert the FAbs into full-length antibodies. The construction of a plasmid encoding the "BS-A" light chain is described here. The other FAbs were cloned using similar procedures. To generate FAb "BS-A" with a signal sequence, a three-step PCR was performed. First, primers 2525-43 (shown below) and 2578-27 (shown below) were used with the FAb cDNA
template. The PCR conditions were: 94 C for 1 min, (94 C for 20 sec., 48 C for 30 sec., 74 C for 30 sec.) for 4 cycles, (94 C for 20 sec., 66 C for 30 sec., 74 C
for 30 sec.) for 25 cycles and 74 C for 5 min. with Pfu polymerase and the appropriate buffer and nucleotides.
The PCR product was then gel purified and then amplified with primers 2148-98 (shown below) and 2578-27 (shown below). Second, primers 2578-26 (shown below) and 2469-67 (shown below) were used again with the FAb cDNA template. The PCR conditions were: 94 C
for 1 min, (94 C for 20 sec., 48 C for 30 sec., 74 C for 30 sec.) for 4 cycles, (94 C for 20 sec., 66 C for 30 sec., 74 C for 30 sec.) for 25 cycles and 74 C for 5 min. with Pfu polymerase and the appropriate buffer and nucleotides. The PCR product was gel isolated and re-amplified using the same conditions. Finally, the gel isolated PCR products were mixed and amplified with primers 2489-36 (shown below) and 2469-67 (shown below). The final PCR product was Qiagen purified, cut with XbaI and Sail, and Qiagen purified. This fragment containing the FAb with a 5' Kozak (translational initiation) site and the following signal sequence for mammalian expression:
MDMRVPAQLLGLLLLWLRGARC (SEQ ID NO:111), was ligated into pDSRa19.
2489-36 5'-C AGC AGA AGC TTC TAG ACC ACC ATG GAC ATG
AGO GTC CCC OCT CAG CTC CTG GG-3' (SEQ ID N0:116) 2525-43 5'-TGG TTG AGA GOT GCC AGA TGT AAT TTT ATG
CTG ACT CAG CCC-3' (SEQ ID NO:117) 2578-27 5'GGC CGC GTA CTT GTT GTT GCT TTG TTT GGA G-3' (SEQ ID NO:118) 2148-98 5'-CC OCT CAG CTC CTG GGG CTC CTG CTA TTG TGG
TTG AGA GGT GCC AGA T-3' (SEQ ID NO:119) 2578-26 5'-AGC AAC ?AC AAG TAC GCG GCC AGC AGC TAC-3' (SEQ ID NO:120) 2469-67 5'-GA AGT CGA CTA TGA ACA TTC TOT AGO AGC-3' (SEQ ID NO:121) Antibody Preparation Expression vectors containing cDNA encoding heavy and light chain full-length antibodies were transfected into CHO cells and cultured under conditions to allow expression of heavy and light chains and secretion into the cell media. The conditioned media was filtered through a 0.45 gm cellulose acetate filter (Corning, Acton, MA) and applied to a Protein G sepharose(Amersham Pharmacia Biotech, Piscataway, NU) column which had been equilibrated with PBS - Dulbecco's Phosphate Buffered Saline without calcium chloride and without magnesium chloride (Gibco BRL Products, Grand Island, NY). After Ar.
.72249-169 sample application the column was washed with PBS until absorbency at 280 nm reached baseline. Elution of protein was achieved using 100 mM Glycine, pH 2.5.
Fractions were collected and immediately neutralized by addition of IM Tris-HC1, pH 9.2. Antibodies were detected by SDS-polyacrylamide gels visualized by Commassie staining.
Fractions containing antibody were pooled, concentrated and diafiltered into PBS using either TM
Centricon 10 (Amicon) or for larger volumes Centriprep 10 (Andcon).
The isolated antibody was characterized by gel filtration on Superose 6 (Amersham Pharmacia .Biotech, Piscataway, NJ) and was shown to run as a monomeric IgG.
Example 6 Affinity measurements of Fab and IgG
The binding constant (Kd), the on rate constant (ka) and off rate constant (kd) were determined by surface plasmon resonance techniques (BIAcore, Pharmacia, Piscataway, New Jersey). BIAcore analysis of Fab and antibody was performed as follows:
The experiments were carried out using BIACORE 20.00 (BIACORE Inc.) at room temperature. CHO expressed hIFNy was immobilized on a CM5 chip. The Fab or Fab IgG at various concentrations were injected over the hu-IFNIir surface. The data was analyzed using BIAEVALUATION 3.1 software (BIACORE, Inc.). The results are shown in Figure 30.
Example 7 Activity measurements of Fab and IgG

BIAcore neutralization assay Neutralization activity of of Fab converted IgGs was tested on BIAcore (see Example 6).
The results are shown in Figure 29. A concentration depedent inhibition of hu IFNy binding to IFNy-R1 with an IC50 of 9 nM was observed for BS-B IgG.
A549 Cell Proliferation assay Neutralization activity of Fab and IgG
measured in A549 proliferation assay (described in Example 1) was performed as follows: A549 cells were treated with a mixture of a targeted Fab or IgG
(various concentrations) and CHO expressed hIFNy (2ng/m1 or 5 ng/m1). Fab concentrations ranged from 0.3-150 g/ml. IgG concentrations ranged from 0.1-100 g/ml. Positive control Ab (Pharmingen B27) concentrations ranged from 0.01-5 g/ml. Cells were stained with Alamar Blue 5 days post treatment, and analyzed 4 hours post staining on an FL500 plate reader. The results are shown in Figure 26 for BS-A
Fab, BS-B Fab and GP-A Fab and in Figure 27 for BS-A
IgG and BS-B IgG. BS-A Fab and BS-B Fab in Figure 26 and BS-A IgG and BS-B IgG in Figure 27 were shown to have neutralization activity, measured as proliferation activity, at high concentrations, about two orders of magnitude higher than the positive control.
While the present invention has been described in terms of preferred embodiments, it is intended that the scope of the claims should not be limited to the embodiments set forth herein, but rather, should be given the broadest interpretation consistent with the specification as a whole.

SEQUENCE LISTING
<110> Amgen, Inc.
<120> Fully Human Antibody Fab Fragments with Human Interferon-Gamma Neutralizing Activity <130> A-799 <140> TED
<141> 17 October 2002 <160> 135 <170> PatentIn version 3.0 <210> 1 <211> 13 <212> PRT
<213> Homo sapiens <400> 1 Thr Gly Ser Ser Gly Ser Ile Ala Ser His Tyr Val Gin <210> 2 <211> 13 <212> PRT
<213> Homo sapiens <400> 2 Thr Gly Ser Ser Gly Ser Ile Ala Ser Asn Tyr Val Gin <210> 3 <211> 13 <212> PRT
<213> Homo sapiens <400> 3 Thr Arg Ser Ser Gly Ser Ile Ala Ser Tyr Tyr Val Gin <210> 4 <211> 16 <212> PRT
<213> Homo sapiens <400> 4 Arg Ala Thr Gin Ser Leu Leu His Gly Asn Gly His Asn Tyr Leu Asp <210> 5 <211> 16 <212> PRT
<213> Homo sapiens <400> 5 Arg Ser Ser Gin Ser Leu Val His Ser Asp Gly Asn Thr Tyr Leu Ser <210> 6 <211> 11 <212> PRT
<213> Homo sapiens <400> 6 Ser Gly Asp Val Leu Ala Arg Lys Tyr Ala Arg <210> 7 <211> 11 <212> PRT
<213> Homo sapiens <400> 7 Gly Gly Asp Asn Leu Gly Gly Lys Ser Leu His <210> 8 <211> 16 <212> PRT
<213> Homo sapiens <400> 8 Arg Ser Ser Gin Ser Leu Leu His Thr Asn Glu Tyr Asn Tyr Leu Asp <210> 9 <211> 13 <212> PRT
<213> Homo sapiens <400> 9 Thr Gly Ser Ser Gly Ser Ile Ala Asn Asn Tyr Val His <210> 10 <211> 12 <212> PRT
<213> Homo sapiens <400> 10 Arg Ala Ser Gin Tyr Val Ser Ser Asn Ser Leu Ala <210> 11 <211> 16 <212> PRT
<213> Homo sapiens <400> 11 Arg Ser Ser Gin Ser Leu Leu Arg Ser Asn Gly Tyr Asn Tyr Leu Ala <210> 12 <211> 7 <212> PRT
<213> Homo sapiens <400> 12 Glu Asp Lys Glu Arg Pro Ser <210> 13 <211> 7 <212> PRT
<213> Homo sapiens <400> 13 Glu Asp Asn Gin Arg Pro Ser <210> 14 <211> 7 <212> PRT
<213> Homo sapiens <400> 14 Glu Asp Asp Gin Arg Pro Ser <210> 15 <211> 7 <212> PRT
<213> Homo sapiens <400> 15 Met Gly Ser Asn Arg Ala Ser <210> 16 <211> 7 <212> PRT
<213> Homo sapiens <400> 16 Lys Ile Ser Asn Arg Phe Ser <210> 17 <211> 7 <212> PRT
<213> Homo sapiens <400> 17 Lys Asp Arg Glu Arg Pro Ser <210> 18 <211> 7 <212> PRT
<213> Homo sapiens <400> 18 Asp Asp Ser Asp Arg Pro Ser <210> 19 <211> 7 <212> PRT
<213> Homo sapiens <400> 19 Leu Gly Ser Asn Arg Ala Pro <210> 20 <211> 7 <212> PRT
<213> Homo sapiens <400> 20 Glu Asp Asp Gln Arg Pro Ser <210> 21 <211> 7 <212> PRT
<213> Homo sapiens <400> 21 Gly Ala Ser Asn Arg Ala Thr <210> 22 <211> 7 <212> PRT
<213> Homo sapiens <400> 22 Leu Ala Ser Asn Arg Ala Ser <210> 23 <211> 10 <212> PRT
<213> Homo sapiens <400> 23 Gin Ser Tyr Asp Ser Ser Asn Gin Trp Val <210> 24 <211> 9 <212> PRT
<213> Homo sapiens <400> 24 Gin Ser Tyr Asp Gly Ser Ala Trp Val <210> 25 <211> 9 <212> PRT .
<213> Homo sapiens <400> 25 Gin Ser Tyr Asp Arg Asn Ser Leu Val <210> 26 <211> 9 <212> PRT
<213> Homo sapiens <400> 26 Met Gin Ala Leu Gin Leu Pro Pro Thr <210> 27 <211> 9 <212> PRT
<213> Homo sapiens <400> 27 Met Gin Ala Thr Gin Leu Pro Tyr Thr <210> 28 <211> 9 <212> PRT
<213> Homo sapiens <400> 28 Tyr Ser Ala Ala Asp Asn Arg Gly Val <210> 29 <211> 11 <212> PRT
<213> Homo sapiens <400> 29 Gin Val Trp Asp Gly Ser Ser Asp Gin Arg Val <210> 30 <211> 9 <212> PRT
<213> Homo sapiens <400> 30 Met Gin Ala Leu Gin Thr Pro Arg Thr <210> 31 <211> 11 <212> PRT
<213> Homo sapiens <400> 31 Gin Ser Tyr Asp Asn Ser Asn Ser Phe Val Val <210> 32 <211> 9 <212> PRT
<213> Homo sapiens <400> 32 Gin Gin Tyr Gly Ser Ser Pro Ile Thr <210> 33 <211> 9 <212> PRT
<213> Homo sapiens <400> 33 Val His Gly Val His Ile Pro Tyr Thr <210> 34 <211> 5 <212> PRT
<213> Homo sapiens <400> 34 Gly Tyr Tyr Trp Ser <210> 35 <211> 5 <212> PRT
<213> Homo sapiens <400> 35 Ser Tyr Ala Met Ser <210> 36 <211> 5 <212> PRT
<213> Homo sapiens <400> 36 Gly Tyr Tyr Trp Ser <210> 37 <211> 7 <212> PRT
<213> Homo sapiens <400> 37 Asn Ala Arg Met Gly Val Ser <210> 38 <211> 5 <212> PRT
<213> Homo sapiens <400> 38 Ser Tyr Ala Met His <210> 39 <211> 5 <212> PRT
<213> Homo sapiens <400> 39 Ser Tyr Ser Met Asn <210> 40 <211> 5 <212> PRT
<213> Homo sapiens <400> 40 Gly Tyr Tyr Trp Ser <210> 41 <211> 7 <212> PRT
<213> Homo sapiens <400> 41 Ser Gly Gly Tyr Ser Trp Ser <210> 42 <211> 5 <212> PRT
<213> Homo sapiens <400> 42 Ser Asn Tyr Met Ser <210> 43 <211> 7 <212> PRT
<213> Homo sapiens <400> 43 Ser Asn Glu Ala Gly Val Gly <210> 44 <211> 16 <212> PRT
<213> Homo sapiens <400> 44 Glu Ile Asn His Ser Gly Ser Thr Asn Tyr Asn Pro Ser Leu Lys Ser <210> 45 <211> 17 <212> PRT
<213> Homo sapiens <400> 45 Ala Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys Gly <210> 46 <211> 16 <212> PRT
<213> Homo sapiens <400> 46 Glu Ile Asn His Ser Gly Ser Thr Asn Tyr Asn Pro Ser Leu Lys Ser <210> 47 <211> 16 <212> PRT
<213> Homo sapiens <400> 47 His Ile Phe Ser Asn Asp Glu Glu Ser Tyr Ser Thr Ser Leu Lys Ser <210> 48 <211> 17 <212> PRT
<213> Homo sapiens <400> 48 Val Ile Ser Tyr Asp Gly Ser Asn Lys Tyr Tyr Ala Asp Ser Val Lys Gly <210> 49 <211> 17 <212> PRT
<213> Homo sapiens <400> 49 Ser Ile Ser Ser Gly Ser Ser Tyr Arg Tyr Asp Ala Asp Ser Val Lys Gly <210> 50 <211> 16 <212> PRT
<213> Homo sapiens , <400> 50 Glu Ile Asn His Ser Gly Ser Thr Asn Tyr Asn Pro Ser Leu Lys Ser <210> 51 <211> 16 <212> PRT
<213> Homo sapiens <400> 51 Tyr Ile Tyr His Ser Gly Ser Thr Tyr Tyr Asn Pro Ser Leu Lys Ser <210> 52 <211> 16 <212> PRT
<213> Homo sapiens <400> 52 Val Ile Tyr Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys Gly <210> 53 <211> 16 <212> PRT

<213> Homo sapiens <400> 53 Leu Leu Tyr Trp Asp Asp Asp Lys Arg Tyr Ser Pro Ser Leu Arg Ser <210> 54 <211> 12 <212> PRT
<213> Homo sapiens <400> 54 Gly Arg Ala Arg Asn Trp Arg Ser Arg Phe Asp Tyr <210> 55 <211> 11 <212> PRT
<213> Homo sapiens <400> 55 Thr Ser Trp Asn Ala Gly Gly Pro Ile Asp Tyr <210> 56 <211> 12 <212> PRT
<213> Homo sapiens <400> 56 Asp Arg Val Gly Tyr Ser Ser Ser Leu Leu Asp Tyr <210> 57 <211> 19 <212> PRT
<213> Homo sapiens <400> 57 Asp Lys Gly Ser Arg Ile Thr Ile Phe Gly Val Val Gly Ser Ala Gly Phe Asp Tyr <210> 58 <211> 9 <212> PRT
<213> Homo sapiens <400> 58 Leu Leu Leu Tyr Glu Gly Phe Asp Pro <210> 59 <211> 15 <212> PRT
<213> Homo sapiens =
<400> 59 Asp Leu Val Leu Thr Met Thr Ser Arg Arg Ala Ala Phe Asp Ile <210> 60 <211> 11 <212> PRT
<213> Homo sapiens <400> 60 Asp Gin Trp Gly Thr Ile Ser Gly Asn Asp Tyr <210> 61 <211> 18 <212> PRT
<213> Homo sapiens <400> 61 Gly Trp Pro Thr Tyr Val Trp Gly Ser Tyr Arg Pro Lys Gly Tyr Phe Asp Tyr , <210> 62 <211> 8 <212> PRT
<213> Homo sapiens <400> 62 Gly Asp Trp Gly Tyr Phe Asp Tyr <210> 63 <211> 9 <212> PRT
<213> Homo sapiens <400> 63 Asp Ala Asp Gly Gly Asp Tyr Gly Tyr <210> 64 <211> 15 <212> PRT
<213> Homo sapiens <400> 64 Arg Leu Val Arg Tyr Gly Gly Tyr Ser Thr Gly Gly Phe Asp Val <210> 65 <211> 669 <212> DNA
<213> Homo sapiens <220>
<221> CDS
<222> (1)..(669) <400> 65 cag gtg cag ctg cag cag tgg ggc gca gga ctg ttg aag cct tcg gag 48 Gin Val Gin Leu Gin Gin Trp Gly Ala Gly Leu Leu Lys Pro Ser Glu acc ctg tcc ctc acc tgc gct gtc tat ggt ggg tcc ttc agt ggt tac 96 Thr Leu Ser Leu Thr Cys Ala Val Tyr Gly Gly Ser Phe Ser Gly Tyr tac tgg agc tgg atc cgc cag ccc cca ggg aag ggg ctg gag tgg att 144 Tyr Trp Ser Trp Ile Arg Gin Pro Pro Gly Lys Gly Leu Glu Trp Ile ggg gaa atc aat cat agt gga agc acc aac tac aac ccg tcc ctc aag 192 Gly Glu Ile Asn His Ser Gly Ser Thr Asn Tyr Asn Pro Ser Leu Lys agt cga gtc acc ata tca gta gac acg tcc aag aac cag ttc tcc ctg 240 Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn Gin Phe Ser Leu aag ctg agc tct gtg acc gcc gcg gac acg gct gtg tat tac tgt gcg 288 Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys Ala aga ggc cgg gca cgg aac tgg aga tcg cgt ttt gac tac tgg ggc cag 336 Arg Gly Arg Ala Arg Asn Trp Arg Ser Arg Phe Asp Tyr Trp Gly Gin gga acc ctg gtc acc gtc tct agt gcc tcc acc aag ggc cca tcg gtc 384 Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val ttc ccc ctg gca ccc tcc tcc aag agc acc tct ggg ggc aca gcg gcc 432 Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala ctg ggc tgc ctg gtc aag gac tac ttc ccc gaa ccg gtg acg gtg tcg 480 Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser tgg aac tca ggc gcc ctg acc agc ggc gtg cac acc ttc ccg gct gtc 528 Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val cta cag tcc tca gga ctc tac tcc ctc agc agc gtg gtg acc gtg ccc 576 Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro tcc agc agc ttg ggc acc cag acc tac atc tgc aac gtg aat cac aag 624 Ser Ser Ser Leu Gly Thr Gin Thr Tyr Ile Cys Asn Val Asn His Lys ccc agc aac acc aag gtg gac aag aaa gtt gag ccc aaa tct tgt 669 Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys <210> 66 <211> 223 <212> PRT
<213> Homo sapiens <400> 66 Gin Val Gin Leu Gin Gin Trp Gly Ala Gly Leu Leu Lys Pro Ser Glu Thr Leu Ser Leu Thr Cys Ala Val Tyr Gly Gly Ser Phe Ser Gly Tyr Tyr Trp Ser Trp Ile Arg Gin Pro Pro Gly Lys Gly Leu Glu Trp Ile Gly Glu Ile Asn His Ser Gly Ser Thr Asn Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn Gin Phe Ser Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys Ala Arg Gly Arg Ala Arg Asn Trp Arg Ser Arg Phe Asp Tyr Trp Gly Gin Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys <210> 67 <211> 672 <212> DNA
<213> Homo sapiens <220>
<221> CDS
<222> (1)..(672) <400> 67 gag gtg cag ctg gtg gag tot ggg gga ggc ttg gta cag cot ggg ggg 48 Glu Val Gin Leu Val Glu Ser Gly Gly Gly Leu Val Gin Pro Gly Gly too ctg aga ctc too tgt gca goo tot gga ttc acc ttt agc agc tat 96 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr gcc atg agc tgg gtc cgc cag gct cca ggg aag ggg ctg gag tgg gtc 144 Ala Net Ser Trp Val Arg Gin Ala Pro Gly Lys Gly Leu Glu Trp Val tca gct att agt ggt agt ggt ggt ago aca tac tac gca gac tcc gtg 192 Ser Ala Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val aag ggc cgg ttc acc atc tcc aga gac aat tcc aag aac acg ctg tat 240 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr ctg caa atg aac agc ctg aga gcc gag gac acg gcc gta tat tac tgt 288 Leu Gin Net Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys gcg aaa gat cgg gtg ggg tat agc agc ago ctt ctt dac tac tgg ggc 336 Ala Lys Asp Arg Val Gly Tyr Ser Ser Ser Leu Leu Asp Tyr Trp Gly cag gga acc ctg gtc acc gtc tot agt gcc too acc aag ggc cca tcg 384 Gin Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser gtc ttc coo ctg gca coo tcc tcc aag agc acc tot ggg ggc aca gcg 432 Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala goo ctg ggc tgc ctg gtc aag gac tac ttc ccc gaa ccg gtg acg gtg 480 Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val tog tgg aac tca ggc goo ctg acc agc ggc gtg cac acc ttc ccg got 528 Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala gtc cta cag tcc tca gga ctc tac tcc ctc agc agc gtg gtg acc gtg 576 Val Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val ccc tcc agc agc ttg ggc acc cag acc tac atc tgc aac gtg aat cac 624 Pro Ser Ser Ser Leu Gly Thr Gin Thr Tyr Ile Cys Asn Val Asn His aag ccc agc aac acc aag gtg gac aag aaa gtt gag ccc aaa tct tgt 672 Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys <210> 68 <211> 224 <212> PRT
<213> Homo sapiens <400> 68 Glu Val Gin Leu Val Glu Ser Gly Gly Gly Leu Val Gin Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr Ala Met Ser Trp Vel Arg Gin Ala Pro Gly Lys Gly Leu Glu Trp Val Ser Ala Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gin Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Lys Asp Arg Val Gly Tyr Ser Ser Ser Leu Leu Asp Tyr Trp Gly Gin Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser Lau Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys <210> 69 <211> 666 <212> DNA
<213> Homo sapiens <220>
<221> CDS
<222> (1)..(666) <400> 69 cag gtc acc ttg aag gag tct ggt cct gtg ctg gtg aaa ccc aca gag 48 Gin Val Thr Leu Lys Glu Ser Gly Pro Val Leu Val Lys Pro Thr Glu acc ctc acg ctg acc tgc acc gtg tct ggg ttc tca ctc agc aat gct 96 Thr Leu Thr Leu Thr Cys Thr Val Ser Gly Phe Ser Leu Ser Asn Ala aga atg ggt gtg agt tgg atc cgt cag ccc cca ggg aag gcc cg gag 144 Arg Met Gly Val Ser Trp Ile Arg Gin Pro Pro Gly Lys Ala Leu Glu tgg ctt gca cac att ttt tcg aat gac gaa gaa tcc tac agc aca tct 192 Trp Leu Ala His Ile Phe Ser Asn Asp Glu Glu Ser Tyr Ser Thr Ser ctg aag agc agg ctc acc atc tcc aag gac acc tcc caa agc cag gtg 240 Leu Lys Ser Arg Leu Thr Ile Ser Lys Asp Thr Ser Gin Ser Gin Val gtc ctt acc atg acc aac atg gac cct gtg gac aca gcc acg tat tac 288 Val Leu Thr Met Thr Asn Met Asp Pro Val Asp Thr Ala Thr Tyr Tyr tgt gca cgg ctt tta ttg tac gag ggg ttc gac ccc tgg ggc cag gga 336 Cys Ala Arg Leu Leu Leu Tyr Glu Gly Phe Asp Pro Trp Gly Gin Gly aCC ctg gtc acc gtc tct agt gcc tcc acc aag ggc cca tcg gtc ttc 384 Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe ccc ctg gca ccc tcc tcc aag agc acc tct ggg ggc aca gcg gcc ctg 432 Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu ggc tgc ctg gtc aag gac tac ttc ccc gaa ccg gtg acg gtg tcg tgg 480 Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp aac tca ggc gcc ctg acc agc ggc gtg cac acc ttc ccg gct gtc cta 528 Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu cag tcc tca gga ctc tac tcc ctc agc agc gtg gtg acc gtg ccc tcc 576 Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser agc agc ttg ggc acc cag acc tac atc tgc aac gtg aat cac aag ccc 624 Ser Ser Leu Gly Thr Gin Thr Tyr Ile Cys Asn Val Asn His Lys Pro agc aac acc aag gtg gac aag aaa gtt gag ccc aaa tct tgt 666 Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys <210> 70 <211> 222 <212> PRT
<213> Homo sapiens <400> 70 Gin Val Thr Leu Lys Glu Ser Gly Pro Val Leu Val Lys Pro Thr Glu Thr Leu Thr Leu Thr Cys Thr Val Ser Gly Phe Ser Leu Ser Asn Ala Arg Met Gly Val Ser Trp Ile Arg Gin Pro Pro Gly Lys Ala Leu Glu Trp Leu Ala His Ile Phe Ser Asn Asp Glu Glu Ser Tyr Ser Thr Ser Leu Lys Ser Arg Leu Thr Ile Ser Lys Asp Thr Ser Gin Ser Gin Val Val Leu Thr Met Thr Asn Met Asp Pro Val Asp Thr Ala Thr Tyr Tyr Cys Ala Arg Leu Leu Leu Tyr Glu Gly Phe Asp Pro Trp Gly Gin Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys <210> 71 <211> 690 <212> DNA
<213> Homo sapiens <220>
<221> CDS
<222> (1)..(690) <400> 71 cag gtg cag cta cag cag tgg ggc gca gga ctg ttg aag cct tcg gag 48 Gin Val Gin Leu Gin Gin Trp Gly Ala Gly Leu Leu Lys Pro Ser Glu acc ctg tcc ctc acc tgc gct gtc tat ggt ggg tcc ttc agt ggt tac 96 Thr Leu Ser Leu Thr Cys Ala Val Tyr Gly Gly Ser Phe Ser Gly Tyr tac tgg agc tgg atc cgc cag ccc cca ggg aag ggg ctg gag tgg att 144 Tyr Trp Ser Trp Ile Arg Gin Pro Pro Gly Lys Gly Leu Glu Trp Ile ggg gaa atc aat cat agt gga agc acc aac tac aac ccg tcc ctc aag 192 Gly Glu Ile Asn His Ser Gly Ser Thr Asn Tyr Asn Pro Ser Leu Lys agt cga gtc acc ata tca gta gac acg tcc aag aac cag ttc tcc ctg 240 Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn Gin Phe Ser Leu aag ctg agc tct gtg acc gcc gcg gac acg gct gtg tat tac tgt gcg 288 Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys Ala aga gat aag ggc tcc cgt att acg att ttt gga gtg gtt ggg tcc gct 336 Arg Asp Lys Gly Ser Arg Ile Thr Ile Phe Gly Val Val Gly Ser Ala ggc ttt gac tac tgg ggc cag ggc acc ctg gtc acc gtc tct agt gcc 384 Gly Phe Asp Tyr Trp Gly Gin Gly Thr Leu Val Thr Val Ser Ser Ala tcc acc aag ggc cca tog gtc ttc ccc ctg gca ccc tcc tcc aag agc 432 Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser acc tct ggg ggc aca gcg gcc ctg ggc tgc ctg gtc aag gac tac ttc 480 Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe ccc gaa cg gtg acg gtg tcg tgg aac tca ggc gcc ctg acc agc ggc 528 Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly gtg cac acc ttc ccg gct gtc cta cag tcc tca gga ctc tac tcc ctc 576 Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu agc agc gtg gtg acc gtg ccc tcc agc agc ttg ggc acc cag acc tac 624 Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr Tyr atc tgc aac gtg aat cac aag ccc agc aac acc aag gtg gac aag aaa 672 Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys gtt gag ccc aaa tct tgt 690 Val Glu Pro Lys Ser Cys <210> 72 <211> 230 <212> PRT
<213> Homo sapiens <400> 72 Gin Val Gin Leu Gin Gin Trp Gly Ala Gly Leu Leu Lys Pro Ser Glu Thr Leu Ser Leu Thr Cys Ala Val Tyr Gly Gly Ser Phe Ser Gly Tyr Tyr Trp Ser Trp Ile Arg Gin Pro Pro Gly Lys Gly Leu Glu Trp Ile Gly Glu Ile Asn His Ser Gly Ser Thr Asn Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn Gin Phe Ser Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys Ala Arg Asp Lys Gly Ser Arg Ile Thr Ile Phe Gly Val Val Gly Ser Ala Gly Phe Asp Tyr Trp Gly Gin Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys <210> 73 <211> 666 <212> DNA
<213> Homo sapiens <220>
<221> CDS
<222> (1)..(666) <400> 73 gag gtg cag ctg ctg gag tct ggg gga ggc ctg gtc aag cot ggg ggg 48 Glu Val Gin Leu Leu Glu Ser Gly Gly Gly Leu Val Lys Pro Gly Gly tcc ctg aga ctc too tgt gca gcc tot gga ttc acc ttc agt ago tat 96 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr ago atg aac tgg gtc cgc cag gct cca ggg aag ggg ctg gag tgg gtc 144 Ser Met Asn Trp Val Arg Gin Ala Pro Gly Lys Gly Leu Glu Trp Val tca tcc att agt agt ggt agc agt tac aga tac gac gca gac tca gtg 192 Ser Ser Ile Ser Ser Gly Ser Ser Tyr Arg Tyr Asp Ala Asp Ser Val aag ggc cga ttc acc atc tcc aga gac aat tcc aag aac acg ctg tat 240 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr ctg caa atg aat agc ctg aga gcc gag gac acg gcc ata tat tac tgt 288 Leu Gin Net Asn Ser Leu Arg Ala Glu Asp Thr Ala Ile Tyr Tyr Cys gcg gat cag atg ggt aca att agt ggc aat gac tac tgg ggc cag ggc 336 Ala Asp Gin Net Gly Thr Ile Ser Gly Asn Asp Tyr Trp Gly Gin Gly acc ctg gtc acc gtc tct agt gcc tcc acc aag ggc cca tcg gtc ttc 384 Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe ccc ctg gca ccc tcc tcc aag agc acc tct ggg ggc aca gcg gcc ctg 432 Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu ggc tgc ctg gtc aag gac tac ttc ccc gaa ccg gtg acg gtg tcg tgg 480 Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp aac tca ggc gcc ctg acc agc ggc gtg cac acc ttc ccg gct gtc cta 528 Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu cag tcc tca gga ctc tac tcc ctc agc agc gtg gtg acc gtg ccc tcc 576 Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser agc agc ttg ggc acc cag acc tac atc tgc aac gtg aat cac aag ccc 624 Ser Ser Leu Gly Thr Gin Thr Tyr Ile Cys Asn Val Asn His Lys Pro agc aac acc aag gtg gac aag aaa gtt gag ccc aaa tct tgt 666 Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys <210> 74 <211> 222 <212> PRT
<213> Homo sapiens <400> 74 Glu Val Gin Leu Leu Glu Ser Gly Gly Gly Leu Val Lys Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr Ser Met Asn Trp Val Arg Gin Ala Pro Gly Lys Gly Leu Glu Trp Val Ser Ser Ile Ser Ser Gly Ser Ser Tyr Arg Tyr Asp Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Ile Tyr Tyr Cys Ala Asp Gin Met Gly Thr Ile Ser Gly Asn Asp Tyr Trp Gly Gin Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys <210> 75 <211> 681 <212> DNA
<213> Homo sapiens <220>
<221> CDS
<222> (1)..(681) <400> 75 cag gtg cag ctg gtg gag acc ggg gga ggc gtg gtc cag cct ggg agg 48 Gin Val Gin Leu Val Glu Thr Gly Gly Gly Val Val Gin Pro Gly Arg tcc ctg aga ctc tcc tgt gca gcc tct gga ttc acc ttc agt agc tat 96 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr gct atg cac tgg gtc cgc cag gct cca ggc aag ggg ctg gag tgg gtg 144 Ala Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val gca gtt ata tca tat gat gga agc aat aaa tac tac gca gac tcc gtg 192 Ala Val Ile Ser Tyr Asp Gly Ser Asn Lys Tyr Tyr Ala Asp Ser Val aag ggc cga ttc acc atc tcc aga gac aat tcc aag aac acg ctg tat 240 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr ctg caa atg aac agc ctg aga gct gag gac acg gct gtg tat tac tgt 288 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys gcg agc gac cta gtc ctt act atg acc tca cga cgg gct gct ttt gat 336 Ala Ser Asp Leu Val Leu Thr Met Thr Ser Arg Arg Ala Ala Phe Asp atc tgg ggc caa ggg aca atg gtc acc gtc tct agt gcc tcc acc aag 384 Ile Trp Gly Gln Gly Thr Met Val Thr Val Ser Ser Ala Ser Thr Lys ggc cca tcg gtc ttc ccc ctg gca ccc tcc tcc aag agc acc tct ggg 432 Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly ggc aca gcg gcc ctg ggc tgc ctg gtc aag gac tac ttc ccc gaa ccg 480 Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro gtg acg gtg tcg tgg aac tca ggc gcc ctg acc agc ggc gtg cac acc 528 Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr ttc ccg gct gtc cta cag tcc tca gga ctc tac tcc ctc agc agc gtg 576 Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val gtg acc gtg ccc tcc agc agc ttg ggc acc cag acc tac atc tgc aac 624 Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn gtg aat cac aag ccc agc aac acc aag gtg gac aag aaa gtt gag ccc 672 Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro aaa tct tgt 681 Lys Ser Cys =
<210> 76 <211> 227 <212> PRT
<213> Homo sapiens <400> 76 Gln Val Gln Leu Val Glu Thr Gly Gly Gly Val Val Gln Pro Gly Arg Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr Ala Met His Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val Ala Val Ile Ser Tyr Asp Gly Ser Asn Lys Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Ser Asp Leu Val Leu Thr Met Thr Ser Arg Arg Ala Ala Phe Asp Ile Trp Gly Gln Gly Thr Met Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys <210> 77 <211> 660 <212> DNA
<213> Homo sapiens <220>
<221> CDS
<222> (1)..(660) <400> 77 gag gtc cag ctg gtg cag tct ggg gga ggc ttg gtc cag cct ggg ggg 48 Glu Val Gin Leu Val Gin Ser Gly Gly Gly Leu Val Gin Pro Gly Gly tcc ctg aga ctc tcc tgt gca gcc tct gga ttc acc gtc agt agc aac 96 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Val Ser Ser Asn tac atg agc tgg gtc cgc cag gct cca ggg aag ggg ctg gag tgg gtc 144 Tyr Met Ser Trp Val Arg Gin Ala Pro Gly Lys Gly Leu Glu Trp Val tca gtt att tat agc ggt ggt agc aca tac tac gca gac tcc gtg aag 192 Ser Val Ile Tyr Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys ggc aga ttc acc atc tcc aga gac aat tcc aag aac acg ctg tat ctt 240 Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr Leu caa atg aac agc ctg aga gcc gag gac acg gct gtg tat tac tgt gcg 288 Gin Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala aga gat tcg gac ggc ggt gac tat ggc tac tgg ggc cag gga acc ctg 336 Arg Asp Ser Asp Gly Gly Asp Tyr Gly Tyr Trp Gly Gin Gly Thr Leu gtc acc gtc tct agt gcc tcc acc aag ggc cca tcg gtc ttc ccc ctg 384 Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu gca ccc tcc tcc aag agc acc tct ggg ggc aca gcg gcc ctg ggc tgc 432 Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys ctg gtc aag gac tac ttc ccc gaa ccg gtg acg gtg tcg tgg aac tca 480 Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser ggc gcc ctg acc agc ggc gtg cac acc ttc ccg gct gtc cta cag tcc 528 Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gin Ser tca gga ctc tac tcc ctc agc agc gtg gtg acc gtg ccc tcc agc agc 576 Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser ttg ggc acc cag acc tac atc tgc aac gtg aat cac aag ccc agc aac 624 Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn acc aag gtg gac aag aaa gtt gag ccc aaa tct tgt 660 Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys <210> 78 <211> 220 <212> PRT
<213> Homo sapiens <400> 78 Glu Val Gln Leu Val Gln Ser Gly Gly Gly Leu Val Gln Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Val Ser Ser Asn Tyr Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val Ser Val Ile Tyr Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Arg Asp Ser Asp Gly Gly Asp Tyr Gly Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys <210> 79 <211> 663 <212> DNA
<213> Homo sapiens <220>
<221> CDS
<222> (1)..(663) <400> 79 cag gtg cag ctg cag gag tcg ggc cca gga ctg gtg aag cot tog gag 48 Gin Val Gin Leu Gin Glu Ser Gly Pro Gly Leu Val Lys Pro Ser Glu acc ctg too ctc acc tgc gct gtc tot ggt ggc tcc atc agc agt ggt 96 Thr Leu Ser Leu Thr Cys Ala Val Ser Gly Gly Ser Ile Ser Ser Gly ggt tac tcc tgg agc tgg atc cgg cag cca cca ggg aag ggc ctg gag 144 Gly Tyr Ser Trp Ser Trp Ile Arg Gin Pro Pro Gly Lys Gly Leu Glu tgg att ggg tac atc tat cat agt ggg agc acc tac tac aac cog too 192 Trp Ile Gly Tyr Ile Tyr His Ser Gly Ser Thr Tyr Tyr Asn Pro Ser ctc aag agt cga gtc acc ata tca gta gac agg too aag aac cag ttc 240 Leu Lys Ser Arg Val Thr Ile Ser Val Asp Arg Ser Lys Asn Gin Phe too ctg aag ctg agc tot gtg acc gcc gcg gac acg gcc gtg tat tac 288 Ser Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr tgt gcc aga ggg gac tgg ggc tac ttt gac tac tgg ggc cag gga acc 336 Cys Ala Arg Gly Asp Trp Gly Tyr Phe Asp Tyr Trp Gly Gin Gly Thr ctg gtc acc gtc tot agt gcc too acc aag ggc cca tog gtc ttc ccc 384 Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro ctg gca ccc too too aag agc acc tot ggg ggc aca gcg gcc ctg ggc 432 Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly tgc ctg gtc aag gac tac ttc coo gaa cog gtg acg gtg tog tgg aac 480 Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn tca ggc gcc ctg acc agc ggc gtg cac acc ttc ccg gct gtc cta cag 528 Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gin tcc tca gga ctc tac tcc ctc agc agc gtg gtg acc gtg ccc tcc agc 576 Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser agc ttg ggc acc cag acc tac atc tgc aac gtg aat cac aag ccc agc 624 Ser Leu Gly Thr Gin Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser aac acc aag gtg gac aag aaa gtt gag ccc aaa tct tgt 663 Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys <210> 80 <211> 221 <212> PRT
<213> Homo sapiens <400> 80 Gin Val Gin Leu Gin Glu Ser Gly Pro Gly Leu Val Lys Pro Ser Glu Thr Leu Ser Leu Thr Cys Ala Val Ser Gly Gly Ser Ile Ser Ser Gly Gly Tyr Ser Trp Ser Trp Ile Arg Gin Pro Pro Gly Lys Gly Leu Glu Trp Ile Gly Tyr Ile Tyr His Ser Gly Ser Thr Tyr Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Val Asp Arg Ser Lys Asn Gin Phe Ser Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys Ala Arg Gly Asp Trp Gly Tyr Phe Asp Tyr Trp Gly Gin Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys <210> 81 <211> 687 <212> DNA
<213> Homo sapiens <220>
<221> CDS
<222> (1)..(687) <400> 81 gag gtg cag cta cag cag tgg ggc gca gga ctg ttg aag cct tcg gag 48 Glu Val Gin Leu Gin Gin Trp Gly Ala Gly Leu Leu Lys Pro Ser Glu acc ctg tcc ctc acc tgc gct gtc tat ggt ggg tcc ttc agt ggt tac 96 Thr Leu Ser Leu Thr Cys Ala Val Tyr Gly Gly Ser Phe Ser Gly Tyr tac tgg agc tgg atc cgc cag ccc cca ggg aag ggg ctg gag tgg att 144 Tyr Trp Ser Trp Ile Arg Gin Pro Pro Gly Lys Gly Leu Glu Trp Ile ggg gaa atc aat cat agt gga agc acc aac tac aac ccg tcc ctc aag 192 Gly Glu Ile Asn His Ser Gly Ser Thr Asn Tyr Asn Pro Ser Leu Lys agt cga gtc acc ata tca gta gac acg tcc aag aac cag ttc tcc ctg 240 Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn Gin Phe Ser Leu aag ctg agc tct gtg acc gcc gcg gac acg gct gtg tat tac tgt gcg 288 Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys Ala aga ggc tgg ccc act tac gtt tgg ggg agt tat cgt ccc aaa ggc tac 336 Arg Gly Trp Pro Thr Tyr Val Trp Gly Ser Tyr Arg Pro Lys Gly Tyr ttt gac tac tgg ggc cag gga acc ctg gtc acc gtc tct agt gcc tcc 384 Phe Asp Tyr Trp Gly Gin Gly Thr Leu Val Thr Val Ser Ser Ala Ser acc aag ggc cca tcg gtc ttc ccc ctg gca ccc tcc tcc aag agc acc 432 Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr tct ggg ggc aca gcg gcc ctg ggc tgc ctg gtc aag gac tac ttc ccc 480 Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro gaa ccg gtg acg gtg tcg tgg aac tca ggc gcc ctg acc agc ggc gtg 528 Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val cac acc ttc ccg gct gtc cta cag tcc tca gga ctc tac tcc ctc agc 576 His Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser agc gtg gtg acc gtg ccc tcc agc agc ttg ggc acc cag acc tac atc 624 Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr Tyr Ile tgc aac gtg aat cac aag ccc agc aac acc aag gtg gac aag aaa gtt 672 Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val gag ccc aaa tct tgt 687 Glu Pro Lys Ser Cys <210> 82 <211> 229 <212> PRT
<213> Homo sapiens <400> 82 Glu Val Gin Leu Gin Gin Trp Gly Ala Gly Leu Leu Lys Pro Ser Glu Thr Leu Ser Leu Thr Cys Ala Val Tyr Gly Gly Ser Phe Ser Gly Tyr Tyr Trp Ser Trp Ile Arg Gin Pro Pro Gly Lys Gly Leu Glu Trp Ile Gly Glu Ile Asn His Ser Gly Ser Thr Asn Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn Gin Phe Ser Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys Ala Arg Gly Trp Pro Thr Tyr Val Trp Gly Ser Tyr Arg Pro Lys Gly Tyr Phe Asp Tyr Trp Gly Gin Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys <210> 83 <211> 684 <212> DNA
<213> Homo sapiens <220>
<221> CDS
<222> (1)..(684) <400> 83 gcc aat acc ctt gaa gag tct ggt cct acg ctg gtg caa ccg aca cag 48 Ala Asn Thr Leu Glu Glu Ser Gly Pro Thr Leu Val Gln Pro Thr Gin 1 5 10. 15 acc ctc acg ctg acc tgc tcc tac tct ggg ttc tca ctc agc agt aat 96 Thr Leu Thr Leu Thr Cys Ser Tyr Ser Gly Phe Ser Leu Ser Ser Asn gaa gcg ggt gtg ggc tgg atc cgt cag ccc cca gga aag gcc ccg gag 144 Glu Ala Gly Val Gly Trp Ile Arg Gin Pro Pro Gly Lys Ala Pro Glu tgg ctt gca ctt ctt tat tgg gat gat gat aag cgc tac agc ccg tct 192 Trp Leu Ala Leu Leu Tyr Trp Asp Asp Asp Lys Arg Tyr Ser Pro Ser ctg agg agc agg ctc atc gtt aac aag gac acc tcc aaa agc cag gtt 240 Leu Arg Ser Arg Leu Ile Val Asn Lys Asp Thr Ser Lys Ser Gin Val gtc ctt aca atg acc aac atg gac cct gtg gac acg gcc aca tat tac 288 Val Leu Thr Met Thr Asn Met Asp Pro Val Asp Thr Ala Thr Tyr Tyr tgt gca cac aga ctc gtc aga tat ggt ggc tac tca acg ggt ggt ttt 336 Cys Ala His Arg Leu Val Arg Tyr Gly Gly Tyr Ser Thr Gly Gly Phe gat gtc tgg ggc caa ggg acc acg gtc acc gtc tca agc gcc tcc acc 384 Asp Val Trp Gly Gin Gly Thr Thr Val Thr Val Ser Ser Ala Ser Thr aag ggc cca tcg gtc ttc ccc ctg gca ccc tcc tcc aag agc acc tct 432 Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser ggg ggc aca gcg gcc ctg ggc tgc ctg gtc aag gac tac ttc ccc gaa 480 Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu ccg gtg acg gtg tcg tgg aac tca ggc gcc ctg acc agc ggc gtc cac 528 Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His acc ttc ccg gct gtc cta cag tcc tca gga ctc tac tcc ctc agc agc 576 Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser gta gtg acc gtg ccc tcc agc agc ttg ggc acc cag acc tac atc tgc 624 Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr Tyr Ile Cys aac gtg aat cac aag ccc agc aac acc aag gtg gac aag aaa gtt gag 672 Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu ccc aaa tct tgt 684 Pro Lys Ser Cys <210> 84 <211> 228 <212> PRT
<213> Homo sapiens <400> 84 Ala Asn Thr Leu Glu Glu Ser Gly Pro Thr Leu Val Gin Pro Thr Gin Thr Leu Thr Leu Thr Cys Ser Tyr Ser Gly Phe Ser Leu Ser Ser Asn Glu Ala Gly Val Gly Trp Ile Arg Gin Pro Pro Gly Lys Ala Pro Glu Trp Leu Ala Leu Leu Tyr Trp Asp Asp Asp Lys Arg Tyr Ser Pro Ser Leu Arg Ser Arg Leu Ile Val Asn Lys Asp Thr Ser Lys Ser Gin Val Val Leu Thr Met Thr Asn Met Asp Pro Val Asp Thr Ala Thr Tyr Tyr Cys Ala His Arg Leu Val Arg Tyr Gly Gly Tyr Ser Thr Gly Gly Phe Asp Val Trp Gly Gin Gly Thr Thr Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys <210> 85 <211> 669 <212> DNA
<213> Homo sapiens <220>
<221> CDS
<222> (1)..(669) <400> 85 gac gtg cag ctg gtg gag act ggg gga ggc ttg gta cag cct ggg ggg 48 Asp Val Gin Leu Val Glu Thr Gly Gly Gly Leu Val Gin Pro Gly Gly tcc ctg aga ctc tcc tgt gcg gcc tct gga ttc acc ttt agc agc tat 96 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr gcc atg agc tgg gtc cgc cag gct cca ggg aag ggg ctg gag tgg gtc 144 Ala Met Ser Trp Val Arg Gin Ala Pro Gly Lys Gly Leu Glu Trp Val tca gct att agt ggt agt ggt ggt agc aca tac tac gca gac tcc gtg 192 Ser Ala Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val aag ggc cgg ttc acc atc tcc aga gac aat tcc aag aac acg ctg tat 240 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr ctg caa atg gac agc ctg aga gcc gag gac acg gcc gta tat tac tgt 288 Leu Gin Met Asp Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys gcg aag acg tcc tgg aac gca ggt ggc ccg att gac tac tgg ggc cag 336 Ala Lys Thr Ser Trp Asn Ala Gly Gly Pro Ile Asp Tyr Trp Gly Gln gga aac ctg gtc acc gtc tca agc gcc tcc acc aag ggc cca tcg gtc 384 Gly Asn Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val ttc ccc ctg gca ccc tcc tcc aag agc acc tct ggg ggc aca gcg gcc 432 Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala ctg ggc tgc ctg gtc aag gac tac ttc ccc gaa ccg gtg acg gtg tcg 480 Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser tgg aac tca ggc gcc ctg acc agc ggc gtc cac acc ttc ccg gct gtc 528 Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val cta cag tcc tca gga ctc tac tcc ctc agc agc gta gtg acc gtg ccc 576 Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro tcc agc agc ttg ggc acc cag acc tac atc tgc aac gtg aat cac aag 624 Ser Ser Ser Leu Gly Thr Gin Thr Tyr Ile Cys Asn Val Asn His Lys ccc agc aac acc aag gtg gac aag aaa gtt gag ccc aaa tct tgt 669 Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys <210> 86 <211> 223 <212> PRT
, <213> Homo sapiens <400> 86 Asp Val Gln Leu Val Glu Thr Gly Gly Gly Leu Val Gln Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Tyr Ala Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val Ser Ala Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gln Met Asp Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Lys Thr Ser Trp Asn Ala Gly Gly Pro Ile Asp Tyr Trp Gly Gln Gly Asn Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser Cys <210> 87 <211> 651 <212> DNA
<213> Homo sapiens <220>
<221> CDS
<222> (1)..(651) <400> 87 aat ttt atg ctg act cag ccc cac tct gtg tcg gag tct ccg ggg aag 48 Asn Phe Met Leu Thr Gin Pro His Ser Val Ser Glu Ser Pro Gly Lys acg gta acc atc tcc tgc acc ggc agc agt ggc agc att gcc agc cac 96 Thr Val Thr Ile Ser Cys Thr Gly Ser Ser Gly Ser Ile Ala Ser His tat gtg cag tgg tac cag cag cgc ccg ggc agt gcc ccc act aat gtg 144 Tyr Val Gin Trp Tyr Gin Gin Arg Pro Gly Ser Ala Pro Thr Asn Val att tat gag gat aag gaa aga ccc tct ggg gtc cct gat cgg ttc tct 192 Ile Tyr Glu Asp Lys Glu Arg Pro Ser Gly Val Pro Asp Arg Phe Ser ggc tcc atc gac agc tcc acc aac tct gcc tcc ctc acc atc tct gga 240 Gly Ser Ile Asp Ser Ser Thr Asn Ser Ala Ser Leu Thr Ile Ser Gly ctg aag act gag gac gag gct gac tac tat tgt cag tct tat gat agc 288 Leu Lys Thr Glu Asp Glu Ala Asp Tyr Tyr Cys Gin Ser Tyr Asp Ser agc aat cag tgg gtg ttc ggc gga ggg acc aag ctg acc gtc cta ggt 336 Ser Asn Gin Trp Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly cag ccc aag gct gcc ccc tcg gtc act ctg ttc ccg ccc tcc tct gag 384 Gin Pro Lys Ala Ala Pro Ser Val Thr Leu Phe Pro Pro Ser Ser Glu gag ctt caa gcc aac aag gcc aca ctg gtg tgt ctc ata agt gac ttc 432 Glu Leu Gin Ala Asn Lys Ala Thr Leu Val Cys Leu Ile Ser Asp Phe tac ccg gga gcc gtg aca gtg gcc tgg aag gca gat agc agc ccc gtc 480 Tyr Pro Gly Ala Val Thr Val Ala Trp Lys Ala Asp Ser Ser Pro Val aag gcg gga gtg gag acc acc aca ccc tcc aaa caa agc aac aac aag 528 Lys Ala Gly Val Glu Thr Thr Thr Pro Ser Lys Gin Ser Asn Asn Lys tac gcg gcc agc agc tac ctg agc ctg acg cct gag cag tgg aag tcc 576 Tyr Ala Ala Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gin Trp Lys Ser cac aga agc tac agc tgc cag gtc acg cat gaa ggg agc acc gtg gag 624 His Arg Ser Tyr Ser Cys Gin Val Thr His Glu Gly Ser Thr Val Glu aag aca gtg gct cct aca gaa tgt tca 651 Lys Thr Val Ala Pro Thr Glu Cys Ser <210> 88 <211> 217 <212> PRT
<213> Homo sapiens <400> 88 Asn Phe Met Leu Thr Gin Pro His Ser Val Ser Glu Ser Pro Gly Lys Thr Val Thr Ile Ser Cys Thr Gly Ser Ser Gly Ser Ile Ala Ser His Tyr Val Gin Trp Tyr Gin Gin Arg Pro Gly Ser Ala Pro Thr Asn Val Ile Tyr Glu Asp Lys Glu Arg Pro Ser Gly Val Pro Asp Arg Phe Ser Gly Ser Ile Asp Ser Ser Thr Asn Ser Ala Ser Leu Thr Ile Ser Gly Leu Lys Thr Glu Asp Glu Ala Asp Tyr Tyr Cys Gin Ser Tyr Asp Ser Ser Asn Gin Trp Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly Gin Pro Lys Ala Ala Pro Ser Val Thr Leu Phe Pro Pro Ser Ser Glu Glu Leu Gin Ala Asn Lys Ala Thr Leu Val Cys Leu Ile Ser Asp Phe Tyr Pro Gly Ala Val Thr Val Ala Trp Lys Ala Asp Ser Ser Pro Val Lys Ala Gly Val Glu Thr Thr Thr Pro Ser Lys Gin Ser Asn Asn Lys Tyr Ala Ala Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gin Trp Lys Ser His. Arg Ser Tyr Ser Cys Gin Val Thr His Glu Gly Ser Thr Val Glu Lys Thr Val Ala Pro Thr Glu Cys Ser <210> 89 <211> 648 <212> DNA
<213> Homo sapiens <220>
<221> CDS
<222> (1)..(648) <400> 89 aat ttt atg ctg act cag ccc cac tct gtg tcg gag tct ccg ggg aag 48 Asn Phe Met Leu Thr Gin Pro His Ser Val Ser Glu Ser Pro Gly Lys acg gta acc atc tcc tgc acc cgc agc agc ggc agc att gcc agc tac 96 Thr Val Thr Ile Ser Cys Thr Arg Ser Ser Gly Ser Ile Ala Ser Tyr tat gtg cag tgg tac cag cag cgc ccg ggc agt tcc ccc acc act gtg 144 Tyr Val Gin Trp Tyr Gin Gin Arg Pro Gly Ser Ser Pro Thr Thr Val atc tat gaa gat gac caa aga ccc tct ggg gtc cct gat cga ttc tct 192 Ile Tyr Glu Asp Asp Gin Arg Pro Ser Gly Val Pro Asp Arg Phe Ser ggc tcc atc gac agt gcc tcc aac tca gcc tcc ctc acc atc tct ggc 240 Gly Ser Ile Asp Ser Ala Ser Asn Ser Ala Ser Leu Thr Ile Ser Gly ctg cag act gag gac gag gct gac tac tat tgt cag tct tat gac agg 288 Leu Gln Thr Glu Asp Glu Ala Asp Tyr Tyr Cys Gin Ser Tyr Asp Arg aac agt ctg gtg ttc ggc ggg ggg acc aag ctg acc gtc ctg ggt cag 336 Asn Ser Leu Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly Gin ccc aag gct gcc ccc tcg gtc act ctg ttc ccg ccc tcc tct gag gag 384 Pro Lys Ala Ala Pro Ser Val Thr Leu Phe Pro Pro Ser Ser Glu Glu ctt caa gcc aac aag gcc aca ctg gtg tgt ctc ata agt gac ttc tac 432 Leu Gin Ala Asn Lys Ala Thr Leu Val Cys Leu Ile Ser Asp Phe Tyr ccg gga gcc gtg aca gtg gcc tgg aag gca gat agc agc ccc gtc aag 480 Pro Gly Ala Val Thr Val Ala Trp Lys Ala Asp Ser Ser Pro Val Lys gcg gga gtg gag acc acc aca ccc tcc aaa caa agc aac aac aag tac 528 Ala Gly Val Glu Thr Thr Thr Pro Ser Lys Gin Ser Asn Asn Lys Tyr gcg gcc agc agc tac ctg agc ctg acg cct gag cag tgg aag tcc cac 576 Ala Ala Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gin Trp Lys Ser His aaa agc tac agc tgc cag gtc acg cat gaa ggg agc acc gtg gag aag 624 Lys Ser Tyr Ser Cys Gin Val Thr His Glu Gly Ser Thr Val Glu Lys aca gtg gct cct aca gaa tgt tca 648 Thr Val Ala Pro Thr Glu Cys Ser <210> 90 <211> 216 <212> PRT
<213> Homo sapiens <400> 90 Asn Phe Met Leu Thr Gin Pro His Ser Val Ser Glu Ser Pro Gly Lys Thr Val Thr Ile Ser Cys Thr Arg Ser Ser Gly Ser Ile Ala Ser Tyr Tyr Val Gin Trp Tyr Gin Gln Arg Pro Gly Ser Ser Pro Thr Thr Val Ile Tyr Glu Asp Asp Gin Arg Pro Ser Gly Val Pro Asp Arg Phe Ser Gly Ser Ile Asp Ser Ala Ser Asn Ser Ala Ser Leu Thr Ile Ser Gly Leu Gin Thr Glu Asp Glu Ala Asp Tyr Tyr Cys Gin Ser Tyr Asp Arg Asn Ser Leu Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly Gin Pro Lys Ala Ala Pro Ser Val Thr Leu Phe Pro Pro Ser Ser Glu Glu Leu Gin Ala Asn Lys Ala Thr Leu Val Cys Leu Ile Ser Asp Phe Tyr Pro Gly Ala Val Thr Val Ala Trp Lys Ala Asp Ser Ser Pro Val Lys Ala Gly Val Glu Thr Thr Thr Pro Ser Lys Gin Ser Asn Asn Lys Tyr Ala Ala Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gin Trp Lys Ser His Lys Ser Tyr Ser Cys Gin Val Thr His Glu Gly Ser Thr Val Glu Lys Thr Val Ala Pro Thr Glu Cys Ser <210> 91 <211> 657 <212> DNA
<213> Homo sapiens <220>
<221> CDS
<222> (1)..(657) <400> 91 gat att gtg atg acc cac act cca ctc tcc tca cct gtc acc ctt gga 48 Asp Ile Val Met Thr His Thr Pro Leu Ser Ser Pro Val Thr Leu Gly cag ccg gcc tcc atc tcc tgc agg tct agt cag agc ctc gta cac agt 96 Gin Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Leu Val His Ser gat gga aac acc tac ttg agt tgg ctt cac cag agg cca ggc cag cct 144 Asp Gly Asn Thr Tyr Leu Ser Trp Leu His Gin Arg Pro Gly Gin Pro cca aga ctc cta att tat aag att tct aac cgg ttc tct ggg gtc cca 192 Pro Arg Leu Leu Ile Tyr Lys Ile Ser Asn Arg Phe Ser Gly Val Pro gac aga ttc agt ggc agt ggg gca ggg aca gat ttc aca ctg aaa atc 240 Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Lys Ile agc agg gtg gaa gct gag gat gtc ggg ctt tat tac tgc atg caa gct 288 Ser Arg Val Glu Ala Glu Asp Val Gly Leu Tyr Tyr Cys Met Gin Ala aca caa ctt ccg tac act ttt ggc cag ggg acc aag ctg gag atc aaa 336 Thr Gin Leu Pro Tyr Thr Phe Gly Gin Gly Thr Lys Leu Glu Ile Lys cga act gtg gct gca cca tct gtc ttc atc ttc ccg cca tct gat gag 384 Arg Thr Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu cag ttg aaa tct gga act gcc tct gtt gtg tgc ctg ctg aat aac ttc 432 Gin Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe tat ccc aga gag gcc aaa gta cag tgg aag gtg gat aac gcc ctc caa 480 Tyr Pro Arg Glu Ala Lys Val Gin Trp Lys Val Asp Asn Ala Leu Gin tcg ggt aac tcc cag gag agt gtc aca gag cag gac agc aag gac agc 528 Ser Gly Asn Ser Gin Glu Ser Val Thr Glu Gin Asp Ser Lys Asp Ser acc tac agc ctc agc agc acc ctg acg ctg agc aaa gca gac tac gag 576 Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu aaa cac aaa gtc tac gcc tgc gaa gtc acc cat cag ggc ctg agc tcg 624 Lys His Lys Val Tyr Ala Cys Glu Val Thr His Gin Gly Leu Ser Ser ccc gtc aca aag agt ttc aac agg gga gag tgt 657 Pro Val Thr Lys Ser Phe Asn Arg Gly Glu Cys <210> 92 <211> 219 <212> PRT
<213> Homo sapiens <400> 92 Asp Ile Val Met Thr His Thr Pro Leu Ser Ser Pro Val Thr Leu Gly Gin Pro Ala Ser Ile Ser Cys Arg Ser Ser Gin Ser Leu Val His Ser Asp Gly Asn Thr Tyr Leu Ser Trp Leu His Gin Arg Pro Gly Gin Pro Pro Arg Leu Leu Ile Tyr Lys Ile Ser Asn Arg Phe Ser Gly Val Pro Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Lys Ile Ser Arg Val Glu Ala Glu Asp Val Gly Leu Tyr Tyr Cys Met Gin Ala Thr Gin Leu Pro Tyr Thr Phe Gly Gin Gly Thr Lys Leu Glu Ile Lys Arg Thr Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gin Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala Lys Val Gin Trp Lys Val Asp Asn Ala Leu Gin Ser Gly Asn Ser Gin Glu Ser Val Thr Glu Gin Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr Ala Cys Glu Val Thr His Gin Gly Leu Ser Ser Pro Val Thr Lys Ser Phe Asn Arg Gly Glu Cys <210> 93 <211> 657 <212> DNA
<213> Homo sapiens <220>
<221> CDS
<222> (1)..(657) <400> 93 gat gtt gtg atg act cag tct cca ctc tcc ctg ccc gtc acc cct gga 48 Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Pro Gly gag ccg gcc tcc atc tcc tgc agg gca act cag agc ctc ctg cat gga 96 Glu Pro Ala Ser Ile Ser Cys Arg Ala Thr Gin Ser Leu Leu His Gly aat gga cac aac tat ttg gat tgg tac ctg cag aag cca ggg cag tct 144 Asn Gly His Asn Tyr Leu Asp Trp Tyr Leu Gin Lys Pro Gly Gin Ser cca cac ctc ctg atc tat atg ggt tct aat cgg gcc tcc ggg gtc cct 192 Pro His Leu Leu Ile Tyr Met Gly Ser Asn Arg Ala Ser Gly Val Pro ggc agg ttc agt ggc act gaa tca ggc aga aat ttt aca ctg aag atc 240 Gly Arg Phe Ser Gly Thr Glu Ser Gly Arg Asn Phe Thr Leu Lys Ile agc aga gtg gag gct gag gat gtt ggg gtc tat tac tgt atg cag gct 288 Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Met Gin Ala cta caa ctt cct ccg acg ttc ggc caa ggt acc agg gtg gat atc aaa 336 Leu Gin Leu Pro Pro Thr Phe Gly Gin Gly Thr Arg Val Asp Ile Lys cga act gtg gct gca cca tct gtc ttc atc ttc ccg cca tct gat gag 384 Arg Thr Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu cag ttg aaa tct gga act gcc tct gtt gtg tgc ctg ctg aat aac ttc 432 Gin Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe tat ccc aga gag gcc aaa gta cag tgg aag gtg gat aac gcc ctc caa 480 Tyr Pro Arg Glu Ala Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gin tcg ggt aac tcc cag gag agt gtc aca gag cag gac agc aag gac agc 528 Ser Gly Asn Ser Gin Glu Ser Val Thr Glu Gin Asp Ser Lys Asp Ser acc tac agc ctc agc agc acc ctg acg ctg agc aaa gca gac tac gag 576 Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu aaa cac aaa gtc tac gcc tgc gaa gtc acc cat cag ggc ctg agc tcg 624 Lys His Lys Val Tyr Ala Cys Glu Val Thr His Gin Gly Leu Ser Ser ccc gtc aca aag agc ttc aac agg gga gag tgt 657 Pro Val Thr Lys Ser Phe Asn Arg Gly Glu Cys <210> 94 <211> 219 <212> PRT
<213> Homo sapiens <400> 94 Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Pro Gly Glu Pro Ala Ser Ile Ser Cys Arg Ala Thr Gin Ser Leu Leu His Gly Asn Gly His Asn Tyr Leu Asp Trp Tyr Leu Gin Lys Pro Gly Gin Ser Pro His Leu Leu Ile Tyr Met Gly Ser Asn Arg Ala Ser Gly Val Pro Gly Arg Phe Ser Gly Thr Glu Ser Gly Arg Asn Phe Thr Leu Lys Ile Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Met Gin Ala Leu Gin Leu Pro Pro Thr Phe Gly Gin Gly Thr Arg Val Asp Ile Lys Arg Thr Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gin Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala Lys Val Gin Trp Lys Val Asp Asn Ala Leu Gin Ser Gly Asn Ser Gin Glu Ser Val Thr Glu Gin Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr Ala Cys Glu Val Thr His Gin Gly Leu Ser Ser Pro Val Thr Lys Ser Phe Asn Arg Gly Glu Cys <210> 95 <211> 642 <212> DNA
<213> Homo sapiens <220>
<221> CDS
<222> (1)..(642) <400> 95 cag tct gtg ctt acg cag ccg ccc tcg gtg tct gtg gcc cca gga aag 48 Gin Ser Val Leu Thr Gin Pro Pro Ser Val Ser Val Ala Pro Gly Lys acg gcc act att acc tgt ggg gga gac aac ctt gga ggt aaa agt cta 96 Thr Ala Thr Ile Thr Cys Gly Gly Asp Asn Leu Gly Gly Lys Ser Leu cac tgg tac cag cag aag cca ggc cag gcc cct gta ctg gtc gtc tac 144 His Trp Tyr Gin Gin Lys Pro Gly Gin Ala Pro Val Leu Val Val Tyr gat gat agc gac cgg ccc tca ggg atc cct gag cga ttt tct ggc tcc 192 Asp Asp Ser Asp Arg Pro Ser Gly Ile Pro Glu Arg Phe Ser Gly Ser aac tct ggg aac acg gcc acc ctg acc att gat agg gtc gaa gac ggg 240 Asn Ser Gly Asn Thr Ala Thr Leu Thr Ile Asp Arg Val Glu Asp Gly gat gag gcc gac tat tat tgt cag gtg tgg gat ggt agt agt gat caa 288 Asp Glu Ala Asp Tyr Tyr Cys Gin Val Trp Asp Gly Ser Ser Asp Gin cga gtc ttc ggc gga ggg acc agg ctg acc gtc cta ggt cag ccc aag 336 Arg Val Phe Gly Gly Gly Thr Arg Leu Thr Val Leu Gly Gin Pro Lys gct gcc ccc tcg gtc act ctg ttc ccg ccc tcc tct gag gag ctt caa 384 Ala Ala Pro Ser Val Thr Leu Phe Pro Pro Ser Ser Glu Glu Leu Gin gcc aac aag gcc aca ctg gtg tgt ctc ata agt gac ttc tac ccg gga 432 Ala Asn Lys Ala Thr Leu Val Cys Leu Ile Ser Asp Phe Tyr Pro Gly gcc gtg aca gtg gcc tgg aag gca gat agc agc ccc gtc aag gcg gga 480 Ala Val Thr Val Ala Trp Lys Ala Asp Ser Ser Pro Val Lys Ala Gly gtg gag acc acc aca ccc tcc aaa caa agc aac aac aag tac gcg gcc 528 Val Glu Thr Thr Thr Pro Ser Lys Gin Ser Asn Asn Lys Tyr Ala Ala agc agc tat ctg agc ctg acg cct gag cag tgg aag tcc cac aga agc 576 Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gin Trp Lys Ser His Arg Ser tac agc tgc cag gtc acg cat gaa ggg agc acc gtg gag aag aca gtg 624 Tyr Ser Cys Gin Val Thr His Glu Gly Ser Thr Val Glu Lys Thr Val gct cct aca gaa tgt tca 642 Ala Pro Thr Glu Cys Ser <210> 96 <211> 214 <212> PRT
<213> Homo sapiens <400> 96 Gin Ser Val Leu Thr Gin Pro Pro Ser Val Ser Val Ala Pro Gly Lys Thr Ala Thr Ile Thr Cys Gly Gly Asp Asn Leu Gly Gly Lys Ser Leu His Trp Tyr Gin Gin Lys Pro Gly Gin Ala Pro Val Leu Val Val Tyr Asp Asp Ser Asp Arg Pro Ser Gly Ile Pro Glu Arg Phe Ser Gly Ser Asn Ser Gly Asn Thr Ala Thr Leu Thr Ile Asp Arg Val Glu Asp Gly Asp Glu Ala Asp Tyr Tyr Cys Gin Val Trp Asp Gly Ser Ser Asp Gin Arg Val Phe Gly Gly Gly Thr Arg Leu Thr Val Leu Gly Gin Pro Lys Ala Ala Pro Ser Val Thr Leu Phe Pro Pro Ser Ser Glu Glu Leu Gin Ala Asn Lys Ala Thr Leu Val Cys Leu Ile Ser Asp Phe Tyr Pro Gly Ala Val Thr Val Ala Trp Lys Ala Asp Ser Ser Pro Val Lys Ala Gly Val Glu Thr Thr Thr Pro Ser Lys Gin Ser Asn Asn Lys Tyr Ala Ala Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gin Trp Lys Ser His Arg Ser Tyr Ser Cys Gin Val Thr His Glu Gly Ser Thr Val Glu Lys Thr Val Ala Pro Thr Glu Cys Ser <210> 97 <211> 636 <212> DNA
<213> Homo sapiens <220>
<221> CDS
<222> (1)..(636) <400> 97 tcc tat gag ctg act cag cca ccc tct gtg tca gtg tct ccg gga cag 48 Ser Tyr Glu Leu Thr Gin Pro Pro Ser Val Ser Val Ser Pro Gly Gin aca gcc agg atc acc tgc tca gga gat gta ctg gca aga aaa tat gct 96 Thr Ala Arg Ile Thr Cys Ser Gly Asp Val Leu Ala Arg Lys Tyr Ala cgg tgg ttc cag cag aag cca ggc cag gcc cct gtg ctg gtg att tat 144 Arg Trp Phe Gin Gin Lys Pro Gly Gin Ala Pro Val Leu Val Ile Tyr aaa gac cgt gag cgg ccc tca ggg atc cct gag cga ttc tcc ggc tcc 192 Lys Asp Arg Glu Arg Pro Ser Gly Ile Pro Glu Arg Phe Ser Gly Ser acc tca ggg acc aca gtc acc ttg acc atc agc ggg gcc cag gtt gaa 240 Thr Ser Gly Thr Thr Val Thr Leu Thr Ile Ser Gly Ala Gln Val Glu gat gag gct gac tat tac tgt tac tct gcg gct gac aac agg ggg gtg 288 Asp Glu Ala Asp Tyr Tyr Cys Tyr Ser Ala Ala Asp Asn Arg Gly Val ttc ggc gga ggg acc aag ctg acc gtc cta cgt cag ccc aag gct gcc 336 Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Arg Gln Pro Lys Ala Ala ccc tcg gtc act ctg ttc cca ccc tcc tct gag gag ctt caa gcc aac 384 Pro Ser Val Thr Leu Phe Pro Pro Ser Ser Glu Glu Leu Gln Ala Asn aag gcc aca ctg gtg tgt ctc ata agt gac ttc tac ccg gga gcc gtg 432 Lys Ala Thr Leu Val Cys Leu Ile Ser Asp Phe Tyr Pro Gly Ala Val aca gtg gcc tgg aag gca gat agc agt ccc gtc aag gcg gga gtg gag 480 Thr Val Ala Trp Lys Ala Asp Ser Ser Pro Val Lys Ala Gly Val Glu acc acc aca ccc tcc aaa caa agc aac aac aag tac gcg gcc agc agc 528 Thr Thr Thr Pro Ser Lys Gln Ser Asn Asn Lys Tyr Ala Ala Ser Ser tac ctg agc ctg acg cct gag cag tgg aag tcc cac aaa agc tac agc 576 Tyr Leu Ser Leu Thr Pro Glu Gln Trp Lys Ser His Lys Ser Tyr Ser tgc cag gtc acg cat gaa ggg agc acc gtg gag aag aca gtg gct cct 624 Cys Gln Val Thr His Glu Gly Ser Thr Val Glu Lys Thr Val Ala Pro aca gaa tgt tca 636 Thr Glu Cys Ser <210> 98 <211> 212 <212> PRT
<213> Homo sapiens <400> 98 Ser Tyr Glu Leu Thr Gln Pro Pro Ser Val Ser Val Ser Pro Gly Gln Thr Ala Arg Ile Thr Cys Ser Gly Asp Val Leu Ala Arg Lys Tyr Ala Arg Trp Phe Gln Gln Lys Pro Gly Gln Ala Pro Val Leu Val Ile Tyr Lys Asp Arg Glu Arg Pro Ser Gly Ile Pro Glu Arg Phe Ser Gly Ser Thr Ser Gly Thr Thr Val Thr Leu Thr Ile Ser Gly Ala Gln Val Glu Asp Glu Ala Asp Tyr Tyr Cys Tyr Ser Ala Ala Asp Asn Arg Gly Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Arg Gln Pro Lys Ala Ala Pro Ser Val Thr Leu Phe Pro Pro Ser Ser Glu Glu Leu Gln Ala Asn Lys Ala Thr Leu Val Cys Leu Ile Ser Asp Phe Tyr Pro Gly Ala Val Thr Val Ala Trp Lys Ala Asp Ser Ser Pro Val Lys Ala Gly Val Glu Thr Thr Thr Pro Ser Lys Gln Ser Asn Asn Lys Tyr Ala Ala Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gln Trp Lys Ser His Lys Ser Tyr Ser Cys Gln Val Thr His Glu Gly Ser Thr Val Glu Lys Thr Val Ala Pro Thr Glu Cys Ser <210> 99 <211> 645 <212> DNA
<213> Homo sapiens <220>
<221> CDS
<222> (1)..(645) <400> 99 gaa att gtg ctc acg cag tct cca ggc acc ctg tct ttg tct cca ggg 48 Glu Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly gaa aga gcc acc ctc tcc tgc cgg gcc agt cag tat gtt agc agc aac 96 Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Tyr Val Ser Ser Asn tcc tta gcc tgg tac cag cag aaa gct ggc cag gct ccc agg ctc ctc 144 Ser Leu Ala Trp Tyr Gln Gln Lys Ala Gly Gln Ala Pro Arg Leu Leu atc tat ggt gca tcc aac agg gcc act ggc atc cca gac agg ttc agt 192 Ile Tyr Gly Ala Ser Asn Arg Ala Thr Gly Ile Pro Asp Arg Phe Ser ggc agt ggg tct ggg aca gac ttc act ctc acc atc agc aga ctg gag 240 Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Arg Leu Glu cct gaa gat ttt gca gtg tat tac tgt cag cag tat ggt agc tcg ccg 288 Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Tyr Gly Ser Ser Pro atc acc ttc ggc caa ggg aca cga ctg gag att aaa cga act gtg gct 336 Ile Thr Phe Gly Gln Gly Thr Arg Leu Glu Ile Lys Arg Thr Val Ala gca cca tct gtc ttc atc ttc ccg cca tct gat gag cag ttg aaa tct 384 Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gin Leu Lys Ser gga act gcc tct gtt gtg tgc ctg ctg aat aac ttc tat ccc aga gag 432 Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu gcc aaa gta cag tgg aag gtg gat aac gcc ctc caa tcg ggt aac tcc 480 Ala Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser Gly Asn Ser cag gag agt gtc aca gag cag gac agc aag gac agc acc tac agc ctc 528 Gln Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr Tyr Ser Leu agc agc acc ctg acg ctg agc aaa gca gac tac gag aaa cac aaa gtc 576 Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val tac gcc tgc gaa gtc acc cat cag ggc ctg agc tcg ccc gtc aca aag 624 Tyr Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro Val Thr Lys agc ttc aac agg gga gag tgt 645 Ser Phe Asn Arg Gly Glu Cys <210> 100 <211> 215 <212> PRT
<213> Homo sapiens <400> 100 Glu Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gin Tyr Val Ser Ser Asn Ser Leu Ala Trp Tyr Gin Gin Lys Ala Gly Gin Ala Pro Arg Leu Leu Ile Tyr Gly Ala Ser Asn Arg Ala Thr Gly Ile Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Arg Leu Glu Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gin Gin Tyr Gly Ser Ser Pro Ile Thr Phe Gly Gin Gly Thr Arg Leu Glu Ile Lys Arg Thr Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gin Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala Lys Val Gin Trp Lys Val Asp Asn Ala Leu Gin Ser Gly Asn Ser Gin Glu Ser Val Thr Glu Gin Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr Ala Cys Glu Val Thr His Gin Gly Leu Ser Ser Pro Val Thr Lys Ser Phe Asn Arg Gly Glu Cys <210> 101 <211> 654 <212> DNA
<213> Homo sapiens <220>

<221> CDS
<222> (1)..(654) <400> 101 aat= ttt atg ctg act cag ccc cac tct gtg tcg gag tct ccg ggg aag 48 Asn Phe Met Leu Thr Gin Pro His Ser Val Ser Glu Ser Pro Gly Lys acg gta acc atc tcc tgc acc ggc agc agt ggc agc att gcc aac aac 96 Thr Val Thr Ile Ser Cys Thr Gly Ser Ser Gly Ser Ile Ala Asn Asn tat gtt cac tgg tac cag caa cgc ccg ggc agt gcc ccc acc act gtg 144 Tyr Val His Trp Tyr Gin Gin Arg Pro Gly Ser Ala Pro Thr Thr Val atc ttt gag gat gac caa aga ccc tct gga gtc cct gat cgg ttc tct 192 Ile Phe Glu Asp Asp Gin Arg Pro Ser Gly Val Pro Asp Arg Phe Ser ggc tcc gtc gac agc tcc tcc aac tct gcc tcc ctc agc att tct gga 240 Gly Ser Val Asp Ser Ser Ser Asn Ser Ala Ser Leu Ser Ile Ser Gly ctg aag act gag gac gag gct gac tac tac tgt cag tct tat gat aac 288 Leu Lys Thr Glu Asp Glu Ala Asp Tyr Tyr Cys Gin Ser Tyr Asp Asn agc aat tca ttt gtg gtg ttc ggc gga ggg acc aag ctg acc gtc cta 336 Ser Asn Ser Phe Val Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu ggt cag ccc aag gct gcc ccc tcg gtc act ctg ttc ccg ccc tcc tct 384 Gly Gin Pro Lys Ala Ala Pro Ser Val Thr Leu Phe Pro Pro Ser Ser gag gag ctt caa gcc aac aag gcc aca ctg gtg tgt ctc ata agt gac 432 Glu Glu Leu Gin Ala Asn Lys Ala Thr Leu Val Cys Leu Ile Ser Asp ttc tac ccg gga gcc gtg aca gtg gcc tgg aag gca gat agc agc ccc 480 Phe Tyr Pro Gly Ala Val Thr Val Ala Trp Lys Ala Asp Ser Ser Pro gtc aag gcg gga gtg gag acc acc aca ccc tcc aaa caa agc aac aac 528 Val Lys Ala Gly Val Glu Thr Thr Thr Pro Ser Lys Gin Ser Asn Asn aag tac gcg gcc agc agc tac ctg agc ctg acg cct gag cag tgg aag 576 Lys Tyr Ala Ala Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gin Trp Lys tcc cac aaa agc tac agc tgc cag gtc acg cat gaa ggg agc acc gtg 624 Ser His Lys Ser Tyr Ser Cys Gin Val Thr His Glu Gly Ser Thr Val gag aag aca gtg gcc cct aca gaa tgc tct 654 Glu Lys Thr Val Ala Pro Thr Glu Cys Ser <210> 102 <211> 218 <212> PRT
<213> Homo sapiens <400> 102 Asn Phe Net Leu Thr Gin Pro His Ser Val Ser Glu Ser Pro Gly Lys Thr Val Thr Ile Ser Cys Thr Gly Ser Ser Gly Ser Ile Ala Asn Asn Tyr Val His Trp Tyr Gin Gin Arg Pro Gly Ser Ala Pro Thr Thr Val Ile Phe Glu Asp Asp Gin Arg Pro Ser Gly Val Pro Asp Arg Phe Ser Gly Ser Val Asp Ser Ser Ser Asn Ser Ala Ser Leu Ser Ile Ser Gly Leu Lys Thr Glu Asp Glu Ala Asp Tyr Tyr Cys Gin Ser Tyr Asp Asn Ser Asn Ser Phe Val Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly Gin Pro Lys Ala Ala Pro Ser Val Thr Leu Phe Pro Pro Ser Ser Glu Glu Leu Gin Ala Asn Lys Ala Thr Leu Val Cys Leu Ile Ser Asp Phe Tyr Pro Gly Ala Val Thr Val Ala Trp Lys Ala Asp Ser Ser Pro Val Lys Ala Gly Val Glu Thr Thr Thr Pro Ser Lys Gin Ser Asn Asn Lys Tyr Ala Ala Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gin Trp Lys Ser His Lys Ser Tyr Ser Cys Gin Val Thr His Glu Gly Ser Thr Val Glu Lys Thr Val Ala Pro Thr Glu Cys Ser <210> 103 <211> 657 <212> DNA
<213> Homo sapiens <220>
<221> CDS
<222> (1)..(657) <400> 103 gaa att gtg ctg act cag tct cca ctc tcc ctt ccc gtc acc cct gga 48 Glu Ile Val Leu Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Pro Gly gag ccg gcc tcc atc tcc tgc agg tct agt cag agc ctc ctg cat act 96 Glu Pro Ala Ser Ile Ser Cys Arg Ser Ser Gin Ser Leu Leu His Thr aat gaa tac aac tat ttg gat tgg tac ctg cag aag cca ggg cag tct 144 Asn Glu Tyr Asn Tyr Leu Asp Trp Tyr Leu Gin Lys Pro Gly Gin Ser cca cag ctc ctc atc tat ttg ggt tct aat cgg gcc ccc ggg gtc cct 192 Pro Gin Leu Leu Ile Tyr Leu Gly Ser Asn Arg Ala Pro Gly Val Pro gac agg ttc agt ggc agt gga tca ggc aca gat ttt aca ctg aga atc 240 Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Arg Ile agc agg gtg gag gct gac gat gtt ggg gtt tac tac tgc atg caa gct 288 Ser Arg Val Glu Ala Asp Asp Val Gly Val Tyr Tyr Cys Net Gin Ala cta caa act cct cgt act ttt ggc cag ggg acc aag ctg gag atc aaa 336 Leu Gin Thr Pro Arg Thr Phe Gly Gin Gly Thr Lys Leu Glu Ile Lys cga act gtg gct gca cca tct gtc ttc atc ttc ccg cca tct gat gag 384 Arg Thr Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu cag ttg aaa tct gga act gcc tct gtt gtg tgc ctg ctg aat aac ttc 432 Gin Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe tat ccc aga gag gcc aaa gta cag tgg aag gtg gat aac gcc ctc caa 480 Tyr Pro Arg Glu Ala Lys Val Gin Trp Lys Val Asp Asn Ala Leu Gin tcg ggt aac tcc cag gag agt gtc aca gag cag gac agc aag gac agc 528 Ser Gly Asn Ser Gin Glu Ser Val Thr Glu Gin Asp Ser Lys Asp Ser acc tac agc ctc agc agc acc ctg acg ctg agc aaa gca gac tac gag 576 Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu aaa cac aaa gtc tac gcc tgc gaa gtc acc cat cag ggc ctg agc tcg 624 Lys His Lys Val Tyr Ala Cys Glu Val Thr His Gin Gly Leu Ser Ser ccc gtc aca aag agc ttc aac agg gga gag tgt 657 Pro Val Thr Lys Ser Phe Asn Arg Gly Glu Cys <210> 104 <211> 219 <212> PRT
<213> Homo sapiens <400> 104 Glu Ile Val Leu Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Pro Gly Glu Pro Ala Ser Ile Ser Cys Arg Ser Ser Gin Ser Leu Leu His Thr Asn Glu Tyr Asn Tyr Leu Asp Trp Tyr Leu Gin Lys Pro Gly Gin Ser Pro Gin Leu Leu Ile Tyr Leu Gly Ser Asn Arg Ala Pro Gly Val Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Arg Ile Ser Arg Val Glu Ala Asp Asp Val Gly Val Tyr Tyr Cys Met Gin Ala Leu Gin Thr Pro Arg Thr Phe Gly Gin Gly Thr Lys Leu Glu Ile Lys Arg Thr Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gin Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala Lys Val Gin Trp Lys Val Asp Asn Ala Leu Gin Ser Gly Asn Ser Gin Glu Ser Val Thr Glu Gin Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr Ala Cys Glu Val Thr His Gin Gly Leu Ser Ser Pro Val Thr Lys Ser Phe Asn Arg Gly Glu Cys <210> 105 <211> 657 <212> DNA
<213> Homo sapiens <220>
<221> CDS
<222> (1)..(657) , <400> 105 gat att gtg atg acc cac act cca ctc tcc ctg ccc gtc acc cct gga 48 Asp Ile Val Met Thr His Thr Pro Leu Ser Leu Pro Val Thr Pro Gly gag ccg gcc tcc atc tcc tgc agg tcc agt cag agc ctc ctg cgt agt 96 Glu Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Leu Leu Arg Ser aat gga tac aac tat ttg gct tgg tac gtg cag aag cca ggg cag tct 144 Asn Gly Tyr Asn Tyr Leu Ala Trp Tyr Val Gin Lys Pro Gly Gin Ser cca caa ctc ctg atc tac ttg gct tct aat cgg gcc tcc ggg gtc cct 192 Pro Gin Leu Leu Ile Tyr Leu Ala Ser Asn Arg Ala Ser Gly Val Pro gac agg ttt agt ggc agt gga tca ggc aca gat ttt aca ctg aag atc 240 Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile agc agc gtg gag gct gag gat gtt ggg gtg tat tac tgc gtg cat ggt 288 Ser Ser Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Val His Gly gta cac att ccc tac act ttt ggc cag ggg acc aag ctg gag atc aaa 336 Val His Ile Pro Tyr Thr Phe Gly Gin Gly Thr Lys Leu Glu Ile Lys cga act gtg gct gca cca tct gtc ttc atc ttc ccg cca tct gat gag 384 Arg Thr Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu cag ttg aaa tct gga act gcc tct gtt gtg tgc ctg ctg aat aac ttc 432 Gin Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe tat ccc aga gag gcc aaa gta cag tgg aag gtg gat aac gcc ctc caa 480 Tyr Pro Arg Glu Ala Lys Val Gin Trp Lys Val Asp Asn Ala Leu Gin tcg ggt aac tcc cag gag agt gtc aca gag cag gac agc aag gac agc 528 Ser Gly Asn Ser Gin Glu Ser Val Thr Glu Gin Asp Ser Lys Asp Ser acc tac agc ctc agc agc acc ctg acg ctg agc aaa gca gac tac gag 576 Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu aaa cac aaa gtc tac gcc tgc gaa gtc acc cat cag ggc ctg agc tcg 624 Lys His Lys Val Tyr Ala Cys Glu Val Thr His Gin Gly Leu Ser Ser ccc gtc aca aag agc ttc aac agg gga gag tgt 657 Pro Val Thr Lys Ser Phe Asn Arg Gly Glu Cys <210> 106 <211> 219 <212> PRT
<213> Homo sapiens <400> 106 Asp Ile Val Net Thr His Thr Pro Leu Ser Leu Pro Val Thr Pro Gly Glu Pro Ala Ser Ile Ser Cys Arg Ser Ser Gin Ser Leu Leu Arg Ser Asn Gly Tyr Asn Tyr Leu Ala Trp Tyr Val Gin Lys Pro Gly Gin Ser Pro Gin Leu Leu Ile Tyr Leu Ala Ser Asn Arg Ala Ser Gly Val Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile Ser Ser Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Val His Gly Val His Ile Pro Tyr Thr Phe Gly Gin Gly Thr Lys Leu Glu Ile Lys Arg Thr Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala Lys Val Gin Trp Lys Val Asp Asn Ala Leu Gin Ser Gly Asn Ser Gin Glu Ser Val Thr Glu Gin Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr Ala Cys Glu Val Thr His Gin Gly Leu Ser Ser Pro Val Thr Lys Ser Phe Asn Arg Gly Glu Cys <210> 107 <211> 648 <212> DNA
<213> Homo sapiens <220>
<221> CDS
<222> (1)..(648) <400> 107 aat ttt atg ctg act cag ccc cac tct gtg tcg gag tct ccg ggg aag 48 Asn Phe Met Leu Thr Gin Pro His Ser Val Ser Glu Ser Pro Gly Lys acg gta acc atc tee tgc acc ggc agc agt ggc agc att gcc agc aac 96 Thr Val Thr Ile Ser Cys Thr Gly Ser Ser Gly Ser Ile Ala Ser Asn tat gtg cag tgg tae cag cag cgc ccg ggc agt gcc ccc acc act gtg 144 Tyr Val Gin Trp Tyr Gin Gin Arg Pro Gly Ser Ala Pro Thr Thr Val atc tat gag gat aac caa aga ccc tct ggg gtc cct cct cgg ttc tct 192 Ile Tyr Glu Asp Asn Gin Arg Pro Ser Gly Val Pro Pro Arg Phe Ser ggc tee atc gac agg tcc tee aac tct gcc tcc ctc ace atc tee gga 240 Gly Ser Ile Asp Arg Ser Ser Asn Ser Ala Ser Leu Thr Ile Ser Gly ctg aag agt gag gac gag get gac tac tae tgt caa tct tat gat ggc 288 Leu Lys Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Gin Ser Tyr Asp Gly agc get tgg gtg ttc ggc gga ggg acc aag ctg acc gtc eta ggt cag 336 Ser Ala Trp Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly Gin ccc aag gct gcc ccc tcg gtc act ctg ttc cca ccc tee tct gag gag 384 Pro Lys Ala Ala Pro Ser Val Thr Leu Phe Pro Pro Ser Ser Glu Glu ctt caa gcc aac aag gcc aca ctg gtg tgt etc ata agt gac ttc tae 432 Leu Gln Ala Asn Lys Ala Thr Leu Val Cys Leu Ile Ser Asp Phe Tyr ccg gga gcc gtg aca gtg gcc tgg aag gca gat agc agc ccc gtc aag 480 Pro Gly Ala Val Thr Val Ala Trp Lys Ala Asp Ser Ser Pro Val Lys gcg gga gtg gag acc acc gca ccc tcc aaa caa agc aac aac aag tac 528 Ala Gly Val Glu Thr Thr Ala Pro Ser Lys Gln Ser Asn Asn Lys Tyr gcg gcc agc agc tac ctg agc ctg acg cct gag cag tgg aag tcc cac 576 Ala Ala Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gln Trp Lys Ser His aaa agc tac agc tgc cag gtc acg cat gaa ggg agc acc gtg gag aag 624 Lys Ser Tyr Ser Cys Gln Val Thr His Glu Gly Ser Thr Val Glu Lys aca gtg gcc cct gca gaa tgc tct 648 Thr Val Ala Pro Ala Glu Cys Ser <210> 108 <211> 216 <212> PRT
<213> Homo sapiens <400> 108 Asn Phe Met Leu Thr Gln Pro His Ser Val Ser Glu Ser Pro Gly Lys Thr Val Thr Ile Ser Cys Thr Gly Ser Ser Gly Ser Ile Ala Ser Asn Tyr Val Gln Trp Tyr Gln Gln Arg Pro Gly Ser Ala Pro Thr Thr Val Ile Tyr Glu Asp Asn Gln Arg Pro Ser Gly Val Pro Pro Arg Phe Ser Gly Ser Ile Asp Arg Ser Ser Asn Ser Ala Ser Leu Thr Ile Ser Gly Leu Lys Ser Glu Asp Glu Ala Asp Tyr Tyr Cys Gln Ser Tyr Asp Gly Ser Ala Trp Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Gly Gln Pro Lys Ala Ala Pro Ser Val Thr Leu Phe Pro Pro Ser Ser Glu Glu Leu Gin Ala Asn Lys Ala Thr Leu Val Cys Leu Ile Ser Asp Phe Tyr Pro Gly Ala Val Thr Val Ala Trp Lys Ala Asp Ser Ser Pro Val Lys Ala Gly Val Glu Thr Thr Ala Pro Ser Lys Gin Ser Asn Asn Lys Tyr Ala Ala Ser Ser Tyr Leu Ser Leu Thr Pro Glu Gin Trp Lys Ser His Lys Ser Tyr Ser Cys Gin Val Thr His Glu Gly Ser Thr Val Glu Lys Thr Val Ala Pro Ala Glu Cys Ser <210> 109 <211> 18 <212> DNA
<213> Homo sapiens <400> 109 ccgactttgc acctagtt 18 <210> 110 <211> 24 <212> DNA
<213> Homo sapiens <400> 110 tttgtcgtct ttccagacgt tagt 24 <210> 111 <211> 22 <212> PRT
<213> Homo sapiens <400> 111 Met Asp Met Arg Val Pro Ala Gin Leu Leu Gly Leu Leu Leu Leu Trp Leu Arg Gly Ala Arg Cys <210> 112 <211> 53 <212> DNA
<213> Homo sapiens <400> 112 cagcagaagc ttctagacca ccatggacat gagggtcccc gctcagccct ggg 53 <210> 113 <211> 48 <212> DNA
<213> Homo sapiens <400> 113 ccgctcagct cctggggctc ctgctattgt ggttgagagg tgccagat 48 <210> 114 <211> 40 <212> DNA
<213> Homo sapiens <400> 114 gtggttgaga ggtgccagat gtcaggtgca gctgcaggag 40 <210> 115 <211> 29 <212> DNA
<213> Homo sapiens <400> 115 gtggaggcac tagagacggt gaccagggt 29 <210> 116 <211> 54 <212> DNA
<213> Homo sapiens <400> 116 cagcagaagc ttctagacca ccatggacat gagggtcccc gctcagctcc tggg 54 <210> 117 <211> 42 <212> DNA
<213> Homo sapiens <400> 117 tggttgagag gtgccagatg taattttatg ctgactcagc cc 42 <210> 118 <211> 31 <212> DNA
<213> Homo sapiens <400> 118 ggccgcgtac ttgttgttgc tttgtttgga g 31 <210> 119 <211> 48 <212> DNA
<213> Homo sapiens <400> 119 ccgctcagct cctggggctc ctgctattgt ggttgagagg tgccagat 48 <210> 120 <211> 30 <212> DNA
<213> Homo sapiens <400> 120 agcaacaaca agtacgcggc cagcagctac 30 <210> 121 <211> 29 <212> DNA
<213> Homo sapiens <400> 121 gaagtcgact atgaacattc tgtaggagc 29 <210> 122 <211> 102 <212> PRT
<213> Homo sapiens <220>
<221> Misc.
<222> (32)..(32) <223> Unidentifiable <220>
<221> Misc.
<222> (33)..(33) <223> Unidentifiable <220>
<221> Misc.
<222> (56)..(56) <223> Unidentifiable <220>
<221> Misc.
<222> (57)..(57) <223> Unidentifiable <220>
<221> Misc.
<222> (58)..(58) <223> Unidentifiable <400> 122 Gin Val Gin Leu Gin Gin Trp Gly Ala Gly Leu Leu Lys Pro Ser Glu Thr Leu Ser Lou Thr Cys Ala Val Tyr Gly Gly Ser Phe Ser Gly Xaa Xaa Tyr Tyr Trp Ser Trp Ile Arg Gin Pro Pro Gly Lys Gly Leu Glu Trp Ile Gly Glu Ile Asn His Xaa Xaa Xaa Ser Gly Ser Thr Asn Tyr Asn Pro Ser Lou Lys Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn Gin Phe Ser Leu Lys Lou Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys Ala Arg <210> 123 <211> 102 <212> PRT
<213> Homo sapiens <220>
<221> Misc.
<222> (32)..(32) <223> Unidentifiable <220>
<221> Misc.
<222> (33)..(33) <223> Unidentifiable <220>
<221> Misc.
<222> (56)..(56) <223> Unidentifiable <220>
<221> Misc.
<222> (57)..(57) <223> Unidentifiable <400> 123 Glu Val Gin Leu Leu Glu Ser Gly Gly Gly Lou Val Gin Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Xaa Xaa Tyr Ala Met Ser Trp Val Arg Gin Ala Pro Gly Lys Gly Leu Glu Trp Val Ser Ala Ile Ser Gly Xaa Xaa Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gin Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Arg <210> 124 <211> 102 <212> PRT
<213> Homo sapiens <220>
<221> Misc.
<222> (55)..(55) <223> Unidentifiable <220>
<221> Misc.
<222> (56)..(56) <223> Unidentifiable <220>
<221> Misc.
<222> (57)..(57) <223> Unidentifiable <400> 124 Gin Val Thr Leu Lys Glu Ser Gly Pro Val Leu Val Lys Pro Thr Glu Thr Leu Thr Leu Thr Cys Thr Val Ser Gly Phe Ser Leu Ser Asn Ala Arg Met Gly Val Ser Trp Ile Arg Gin Pro Pro Gly Lys Ala Leu Glu Trp Leu Ala His Ile Phe Xaa Xaa Xaa Ser Asn Asp Glu Lys Ser Tyr Ser Thr Ser Leu Lys Ser Arg Leu Thr Ile Ser Lys Asp Thr Ser Lys Ser Gin Val Val Leu Thr Met Thr Asn Met Asp Pro Val Asp Thr Ala Thr Tyr Tyr Cys Ala Arg <210> 125 <211> 102 <212> PRT
<213> Homo sapiens <220>
<221> Misc.
<222> (32)..(32) <223> Unidentified <220>
<221> Misc.
<222> (33)¨(33) <223> Unidentified <220>
<221> Misc.
<222> (56)..(56) <223> Unidentified <220>
<221> Misc.
<222> (57)¨(57) <223> Unidentified <400> 125 Glu Val Gin Leu Val Glu Ser Gly Gly Gly Leu Val Lys Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Xaa Xaa Tyr Ser Met Asn Trp Val Arg Gin Ala Pro Gly Lys Gly Leu Glu Trp Val Ser Ser Ile Ser Ser Xaa Xaa Ser Ser Ser Tyr Ile Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Ser Leu Tyr Leu Gin Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Arg <210> 126 <211> 102 <212> PRT
<213> Homo sapiens <220>
<221> Misc.
<222> (32)..(32) <223> Unidentified <220>
<221> Misc.
<222> (33)¨(53) <223> Unidentified <220>
<221> Misc.
<222> (56)..(56) <223> Unidentified <220>
<221> Misc.
<222> (57)..(57) <223> Unidentified <400> 126 Gin Val Gin Leu Val Glu Ser Gly Gly Gly Val Val Gin Pro Gly Arg Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Xaa Xaa Tyr Ala Met His Trp Val Arg Gin Ala Pro Gly Lys Gly Leu Glu Trp Val Ala Val Ile Ser Tyr Xaa Xaa Asp Gly Ser Asn Lys Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gin Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Arg <210> 127 <211> 102 <212> PRT
<213> Homo sapiens <220>
<221> Misc.
<222> (32)..(32) <223> Unidentified <220>
<221> Misc.
<222> (55)¨(55) <223> Unidentified <220>
<221> Misc.
<222> (55)¨(55) <223> Unidentified <220>
<221> Misc.
<222> (56)..(56) <223> Unidentified <220>
<221> Misc.
<222> (57)..(57) <223> Unidentified <400> 127 Glu Val Gin Leu Val Glu Ser Gly Gly Gly Leu Ile Gin Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Val Ser Ser Xaa Xaa Asn Tyr Met Ser Trp Val Arg Gin Ala Pro Gly Lys Gly Leu Glu Trp Val Ser Val Ile Tyr Xaa Xaa Xaa Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gin Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Arg <210> 128 <211> 102 <212> PRT
<213> HOMD sapiens <220>
<221> Misc.
<222> (55)..(55) <223> Unidentified <220>
<221> Misc.
<222> (56)..(56) <223> Unidentified <220>
<221> Misc.
<222> (57)..(57) <223> Unidentified <400> 128 Gin Val Gin Leu Gin Glu Ser Gly Pro Gly Leu Val Lys Pro Ser Glu Thr Leu Ser Leu Thr Cys Thr Val Ser Gly Gly Ser Val Ser Ser Gly Gly Tyr Tyr Trp Ser Trp Ile Arg Gin Pro Pro Gly Lys Gly Leu Glu Trp Ile Gly Tyr Ile Tyr Xaa Xaa Xaa Tyr Ser Gly Ser Thr Asn Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn Gin Phe Ser Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys Ala Arg <210> 129 <211> 102 <212> PRT
<213> Homo sapiens <220>
<221> Misc.
<222> (55)..(55) <223> Unidentified <220>
<221> Misc.
<222> (56)..(56) <223> Unidentified <220>
<221> Misc.
<222> (57)..(57) <223> Unidentified <400> 129 Gin Ile Thr Leu Lys Glu Ser Gly Pro Thr Leu Val Lys Pro Thr Gin Thr Leu Thr Leu Thr Cys Thr Phe Ser Gly Phe Ser Leu Ser Thr Ser Gly Val Gly Val Gly Trp Ile Arg Gin Pro Pro Gly Lys Ala Leu Glu Trp Leu Ala Leu Ile Tyr Xaa Xaa Xaa Trp Asn Asp Asp Lys Arg Tyr Ser Pro Ser Leu Lys Ser Arg Leu Thr Ile Thr Lys Asp Thr Ser Lys Asn Gln Val Val Leu Thr Met Thr Asn Met Asp Pro Val Asp Thr Ala Thr Tyr Tyr Cys Ala His <210> 130 <211> 103 <212> PRT
<213> Homo sapiens <220>
<221> Misc.
<222> (7)..(7) <223> Unidentified <220>
<221> Misc.
<222> (33)..(33) <223> Unidentified <220>
<221> Misc.
<222> (34)..(34) <223> Unidentified <220>
<221> Misc.
<222> (35)¨(35) <223> Unidentified <220>
<221> Misc.
<222> (36)..(36) <223> Unidentified <400> 130 Asn Phe Met Leu Thr Gln Xaa Pro His Ser Val Ser Glu Ser Pro Gly Lys Thr Val Thr Ile Ser Cys Thr Arg Ser Ser Gly Ser Ile Ala Ser Xaa Xaa Xaa Xaa Asn Tyr Val Gln Trp Tyr Gln Gln Arg Pro Gly Ser Ser Pro Thr Thr Val Ile Tyr Glu Asp Asn Gln Arg Pro Ser Gly Val Pro Asp Arg Phe Ser Gly Ser Ile Asp Ser Ser Ser Asn Ser Ala Ser Leu Thr Ile Ser Gly Leu Lys Thr Glu Asp Glu Ala Asp Tyr Tyr Cys Gin Ser Tyr Asp Ser Ser Asn <210> 131 <211> 101 <212> PRT
<213> Homo sapiens <220>
<221> Misc.
<222> (33)..(33) <223> Unidentified <400> 131 Asp Ile Val Met Thr Gin Thr Pro Leu Ser Ser Pro Val Thr Leu Gly Gin Pro Ala Ser Ile Ser Cys Arg Ser Ser Gin Ser Leu Val His Ser Xaa Asp Gly Asn Thr Tyr Leu Ser Trp Leu Gin Gin Arg Pro Gly Gin Pro Pro Arg Leu Leu Ile Tyr Lys Ile Ser Asn Arg Phe Ser Gly Val Pro Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Lys Ile Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Met Gin Ala Thr Gin Phe Pro <210> 132 <211> 101 <212> PRT
<213> Homo sapiens <220>
<221> Misc.
<222> (33)..(33) <223> Unidentified <400> 132 Asp Ile Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Pro Gly Glu Pro Ala Ser Ile Ser Cys Arg Ser Ser Gin Ser Leu Leu His Ser Xaa Asn Gly Tyr Asn Tyr Leu Asp Trp Tyr Leu Gin Lys Pro Gly Gin Ser Pro Gin Leu Leu Ile Tyr Leu Gly Ser Asn Arg Ala Ser Gly Val Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Met Gin Ala Leu Gin Thr Pro <210> 133 <211> 106 <212> PRT
<213> Homo sapiens <220>
<221> Misc.
<222> (10)..(10) <223> Unidentified <220>
<221> Misc.
<222> (26)..(26) <223> Unidentified <220>
<221> Misc.
<222> (29)..(29) <223> Unidentified <220>
<221> Misc.
<222> (34)..(34) <223> Unidentified <220>
<221> Misc.
<222> (55)..(55) <223> Unidentified <220>
<221> Misc.
<222> (56)..(56) <223> Unidentified <220>
<221> Misc.
<222> (57)..(57) <223> Unidentified <220>
<221> Misc.

<222> (58)..(58) <223> Unidentified <220>
<221> Misc.
<222> (76)..(76) <223> Unidentified <220>
<221> Misc.
<222> (77)..(77) <223> Unidentified <400> 133 Ser Tyr Val Leu Thr Gin Pro Pro Ser Xaa Val Ser Val Ala Pro Gly Lys Thr Ala Arg Ile Thr Cys Gly Gly Xaa Asn Asn Xaa Ile Gly Ser Lys Xaa Ser Val His Trp Tyr Gin Gin Lys Pro Gly Gin Ala Pro Val Leu Val Val Tyr Asp Asp Xaa Xaa Xaa Xaa Ser Asp Arg Pro Ser Gly Ile Pro Glu Arg Phe Ser Gly Ser Asn Ser Gly Xaa Xaa Asn Thr Ala Thr Leu Thr Ile Ser Arg Val Glu Ala Gly Asp Glu Ala Asp Tyr Tyr Cys Gin Val Trp Asp Ser Ser Ser Asp His <210> 134 <211> 104 <212> PRT
<213> Homo sapiens <220>
<221> Misc.
<222> (10)..(10) <223> Unidentified <220>
<221> Misc.
<222> (26)..(26) <223> Unidentified <220>
<221> Misc.
<222> (29)..(29) <223> Unidentified <220>
<221> Misc.
<222> (34)..(34) <223> Unidentified <220>
<221> Misc.
<222> (55)..(55) <223> Unidentified <220>
<221> Misc.
<222> (56)..(56) <223> Unidentified <220>
<221> Misc.
<222> (57)..(57) <223> Unidentified <220>
<221> Misc.
<222> (58)..(58) <223> Unidentified <220>
<221> Misc.
<222> (76)..(76) <223> Unidentified <220>
<221> Misc.
<222> (77)..(77) <223> Unidentified <400> 134 Ser Tyr Glu Leu Thr Gin Pro Ser Ser Xaa Val Ser Val Ser Pro Gly Gin Thr Ala Arg Ile Thr Cys Ser Gly Xaa Asp Val Xaa Leu Ala Lys Lys Xaa Tyr Ala Arg Trp Phe Gin Gin Lys Pro Gly Gin Ala Pro Val Leu Val Ile Tyr Lys Asp Xaa Xaa Xaa Xaa Ser Glu Arg Pro Ser Gly Ile Pro Glu Arg Phe Ser Gly Ser Ser Ser Gly Xaa Xaa Thr Thr Val Thr Leu Thr Ile Ser Gly Ala Gin Val Glu Asp Glu Ala Asp Tyr Tyr Cys Tyr Ser Ala Ala Asp Asn Asn <210> 135 <211> 101 <212> PRT
<213> Homo sapiens <220>
<221> Misc.
<222> (33)..(33) <223> Unidentified <220>
<221> Misc.
<222> (34)..(34) <223> Unidentified <220>
<221> Misc.
<222> (35)..(35) <223> Unidentified <220>
<221> Misc.
<222> (36)..(36) <223> Unidentified <220>
<221> Misc.
<222> (37)..(37) <223> Unidentified <400> 135 Glu Ile Val Leu Thr Gin Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gin Ser Val Ser Ser Ser Xaa Xaa Xaa Xaa Xaa Tyr Leu Ala Trp Tyr Gln Gin Lys Pro Gly Gin Ala Pro Arg Leu Leu Ile Tyr Gly Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Arg Leu Glu Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gin Gin Tyr Gly Ser Ser Pro

Claims

CLAIMS:

1. An isolated human antibody that binds specifically to human interferon gamma, wherein the antibody comprises:
(a) a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:34, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:44, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:54, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:1, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:12, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:23;
(b) a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:35, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:45, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:55, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:2, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:13, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:24;
(c) a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:35, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:45, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:56, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:3, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:14, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:25;
(d) a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:36, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:46, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:57, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:4, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:15, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:26;
(e) a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:37, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:47, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:58, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:5, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:16, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:27;
(f) a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:38, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:48, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:59, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:6, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:17, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:28;
(g) a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:39, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:49, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:60, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:7, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:18, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:29;
(h) a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:40, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:50, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:61, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:8, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:19, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:30;
(i) a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:41, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:51, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:62, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:9, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:20, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:31;
(j) a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:42, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:52, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:63, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:10, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:21, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:32; or (k) a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:43, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:53, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:64, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:11, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:22, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:33.

2. The antibody of claim 1, wherein the antibody comprises a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:34, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:44, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:54, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:1, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:12, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:23.

3. The antibody of claim 1, wherein the antibody comprises a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:35, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:45, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:55, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:2, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:13, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:24.

4. The antibody of claim 1, wherein the antibody comprises a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:35, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:45, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:56, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:3, a light chain CDR2 consisting of the amino acid sequence of SEQ ID

NO:14, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:25.

5. The antibody of claim 1, wherein the antibody comprises a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:36, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:46, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:57, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:4, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:15, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:26.

6. The antibody of claim 1, wherein the antibody comprises a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:37, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:47, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:58, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:5, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:16, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:27.

7. The antibody of claim 1, wherein the antibody comprises a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:38, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:48, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:59, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:6, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:17, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:28.

8. The antibody of claim 1, wherein the antibody comprises a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:39, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:49, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:60, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:7, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:18, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:29.

9. The antibody of claim 1, wherein the antibody comprises a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:40, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:50, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:61, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:8, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:19, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:30.

10. The antibody of claim 1, wherein the antibody comprises a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:41, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:51, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:62, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:9, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:20, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:31.

11. The antibody of claim 1, wherein the antibody comprises a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:42, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:52, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:63, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:10, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:21, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:32.

12. The antibody of claim 1, wherein the antibody comprises a heavy chain complementarity determining region 1 (CDR1) consisting of the amino acid sequence of SEQ ID NO:43, a heavy chain complementarity region 2 (CDR2) consisting of the amino acid sequence of SEQ ID NO:53, a heavy chain complementarity determining region 3 (CDR3) consisting of the amino acid sequence of SEQ ID NO:64, a light chain CDR1 consisting of the amino acid sequence of SEQ ID NO:11, a light chain CDR2 consisting of the amino acid sequence of SEQ ID
NO:22, and a light chain CDR3 consisting of the amino acid sequence of SEQ ID NO:33.

13. The antibody of any one of claims 1, 2, 5, 9, and 10, wherein the antibody comprises a heavy chain comprising an amino acid sequence at least 85% identical to SEQ ID NO:66.

14. The antibody of any one of claims 1, 2, 5, 9 and 10 wherein the antibody comprises a heavy chain comprising an amino acid sequence at least 90% identical to SEQ ID NO:66.

15. The antibody of any one of claims 1, 3, 4, and 11, wherein the antibody comprises a heavy chain comprising an amino acid sequence at least 90% identical to SEQ ID NO:68.

16. The antibody of any one of claims 1, 2, 5, 9, and 10, wherein the antibody comprises a heavy chain comprising an amino acid sequence at least 85% identical to SEQ ID NO:72.

17. The antibody of any one of claims 1, 2, 5, 9 and 10 wherein the antibody comprises a heavy chain comprising an amino acid sequence at least 90% identical to SEQ ID NO:72.

18. The antibody of any one of claims 1, 2, 5, 9, and 10, wherein the antibody comprises a heavy chain comprising an amino acid sequence at least 90% identical to SEQ ID NO:82.

19. The antibody of any one of claims 1, 2, 5, 9, and 10, wherein the antibody comprises a heavy chain comprising an amino acid sequence at least 85% identical to SEQ ID NO:80.

20. The antibody of any one of claims 1, 3, 4, and 11, wherein the antibody comprises a heavy chain comprising an amino acid sequence at least 85% identical to SEQ ID NO:78.

21. The antibody of any one of claims 1, 3, 4, and 11, wherein the antibody comprises a heavy chain comprising an amino acid sequence at least 85% identical to SEQ ID NO:86.

22. The antibody of claim 1 or 13, wherein the antibody comprises a light chain comprising an amino acid sequence at least 85% identical to SEQ ID NO:88.

23. The antibody of claim 1 or 15, wherein the antibody comprises a light chain comprising an amino acid sequence at least 85% identical to SEQ ID NO:90.

24. The antibody of claim 1 or 16, wherein the antibody comprises a light chain comprising an amino acid sequence at least 85% identical to SEQ ID NO:94.

25. The antibody of claim 1 or 19, wherein the antibody comprises a light chain comprising an amino acid sequence at least 85% identical to SEQ ID NO:102.

26. The antibody of claim 1 or 21, wherein the antibody comprises a light chain comprising an amino acid sequence at least 85% identical to SEQ ID NO:108.

27. The antibody of claim 1, wherein the antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO:66 and a light chain comprising the amino acid sequence of SEQ ID NO:88.

28. The antibody of claim 1, wherein the antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO:68 and a light chain comprising the amino acid sequence of SEQ ID NO:90.

29. The antibody of claim 1, wherein the antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO:70 and a light chain comprising the amino acid sequence of SEQ ID NO:92.

30. The antibody of claim 1, wherein the antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO:72 and a light chain comprising the amino acid sequence of SEQ ID NO:94.

31. The antibody of claim 1, wherein the antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO:74 and a light chain comprising the amino acid sequence of SEQ ID NO:96.

32. The antibody of claim 1, wherein the antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO:76 and a light chain comprising the amino acid sequence of SEQ ID NO:98.

33. The antibody of claim 1, wherein the antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO:78 and a light chain comprising the amino acid sequence of SEQ ID NO:100.

34. The antibody of claim 1, wherein the antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO:80 and a light chain comprising the amino acid sequence of SEQ ID NO:102.

35. The antibody of claim 1, wherein the antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO:82 and a light chain comprising the amino acid sequence of SEQ ID NO:104.

36. The antibody of claim 1, wherein the antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO:84 and a light chain comprising the amino acid sequence of SEQ ID NO:106.

37. The antibody of claim 1, wherein the antibody comprises a heavy chain comprising the amino acid sequence of SEQ ID NO:86 and a light chain comprising the amino acid sequence of SEQ ID NO:108.

38. The antibody of any one of claims 1 to 37, wherein the isotype of the antibody is IgG.

39. A nucleic acid molecule encoding the antibody of any one of claims 1 to 38.

40. A pharmaceutical composition comprising the antibody of any one of claims 1 to 38 and a pharmaceutically acceptable carrier.

41. Use of a therapeutically effective amount of the antibody of any one of claims 1 to 38 in the manufacture of a medicament for preventing or treating an autoimmune disease or an inflammatory condition.

42. Use of a therapeutically effective amount of the antibody of any one of claims 1 to 38 for preventing or treating an autoimmune disease or an inflammatory condition.

43. The use of claim 41 or 42, wherein the autoimmune disease or inflammatory condition is selected from the group consisting of rheumatoid arthritis, juvenile rheumatoid arthritis, inflammatory bowel disease, and systemic lupus erythematosus.