Title of Invention	AN ANTIBODY OR ANTIGEN BINDING FRAGMENT AND A PHARMACEUTICAL COMPOSITION COMPRISING THE SAME
Abstract	The present invention realates to an antibody or antigen binding fragment thereof that specifically binds to P-Selectin Glycoprotein Ligand-l (PSGL-I) on the surface of an activated T cell and induces apoptosis of the activated T cell for use as a medicament. This 'invention also relates to a pharmaceutical composition comprising the same.

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MODULATORS Off P-SELECTIN GLYCOPROTEIN LIGAND1 Related Application This application clain:s priority from U.S. provisional application serial number: tr 60/310,196, filed on August 3.2001, the content of v/hich is incorporated herd;] by reference. Held of tliejiivcntion The invention relates to compositions and methods for controlling immune responds. Jiackgiound of the Invention The control of unwanted immune responses i$ \ critical issue in the treatment cf divi^35d.:3 such as autoimmune diseases, transplant rejection, allergic diseases, and some ca-'K'.^-ij, The activity of overly aggressive T cells can be controlled by ur.mi'.nosuppression or by die induction of immunological tolerance, Tolerance is dot™*:, as a state where the immune system is made unresponsive to an antigen, whereas immunosuppression, which decreases the immune response :o antigens, usuji.l!.y requires the continued use of medication In organ transplantation, T cells play ar. e1 Sfrmtial role in the immune response to alloaatigsns. Current immunosuppressive r^hres commonly involve the use of cot ticosteroid, cyclosporin or rapamycin, which bloi.k \hz transcription of IL-2, a key gro'.vdi factor for T cells, or inhibit IL-2 dependent proliferation. H:>wever» a number of monoclonal antibodies which either act asi T cell-depleting agents (e.g. C.D3, CD4, CDS)., or as inhibitors of'the cytokine aigrtiil ing or the c\>stimu]a1:oj:y pathways of T calls (e.g. CD25, B7-1, B7-2, CD152, CT'L/k.4) have demonstrated effectiveness in reducing the incidence of rejection with limitsd tide effects or toxicity. Some of these agents nave been shown to have some decree of success in treating autoimmune disease and in prolonging graft survival. Apoptosis is widely believed to he of vital importance for maintaining the proper function of the immune system and a major mechanism to remove unwanted cdis (Kabelite ei al Immunol. Today 14:338-340 (1993); Raff, Natu«:356:397-399 C! 99?.)). Various signals originating from either inside or outside a ceU influence the iifij. a;ad death of tiie ceil. Antibodies against'! :-ell surface molecules such as Fas (or CDW, MW = 43 kD), TNJFR2 (MW = 75 KD), CD2 (MW - 45 KD) and CTLA-4 (MW sa 3: kD) tc induce the apoptosis of T cells (Gsbonie, Cut:. Opin. Immunol, 8:245-248 (lPJift); Lin et a}. J. Immunol. 158:598-603 (1997); Zhang et al. Naturfc:377:348-35Q (ltttf); Lai & al. Eur. J. Immunol 25:3243-3248 (199:1); Mollereau at al. J. Immunol. 156;:?i:?4-3190 (1996); Gribixm et al. Proe. Natl. Acad. Sri. USA 92:&il-815 (1995)). Attempts to use Fas and TNFR2 molecules to control unwantedT cells have been hampeied by the fact chat these two molecules are expressed not only on immune cells, bin: al:;i! on several other important organ systems like liver. This expression pattern pot-; r:i;;illy limits the therapeutic application of these two antibodies (Ogasawara et ah N3.1i're 364:306-809 (1993); Pfeffer et al. Cell:73:457-467 (1993); Engelmann et al. J. Bi:-I ogfcal Chemistry 265:14497-14504 (1990)), Summary, of the Invention The invention is based on the discovery that T cells can be depleted and/or in.:.ucsd to undergo apoptosis by the engagement of the T ceil surface antigen P-ScOtt'tin Glycoprotein Ligand-1 (PSGL-1). T cell depletion can be particularly useful foi "is. treatment of condition* associated with an excessive or unwanted T cell-muliai:ed immune response or excessive or unwanted T cell proliferation. For example. tin;; depletion of T cells can cause the reduction or elimination of uudesirable T cell activity or proliferation associated with autoimmune diseases, transplant rejection, al!:er,gie diseases, and/or T cdl-derived cancers. The invention encompasses methods c:; using tacdulators of PSGL-1 function to prevent or reduce a T cell-mediated i:.-tYi!QUue. response as well as methods of screening for modulators of PSGL-1 function. In one aspect, the invention features a method of preventing or reducing a T C'*li-:n lediated inanune response in an individual. The method includes the following &■:*;).;.: selecting an individual diagnosed as having or as being at risk of acquiring a condi lion characterized by an excessive or unwanted T cell-mediated immune response; ami 3:limnistermg to the individual a compound that binds to PSGL-1 on the surface of ;i T :::iU, wherein the binding of the compound to PSGL-1 on the surface of the T cell i ;•: duces a signal transduction pathway that results in the death o; this T cell, thereby :*;:•: anting or reducing a T cell-mediated immune response in the individual. The compound used in such a method can include an antibody or antigen fori utiing fragment thereof that specifically binds to PSGL-1. In one example, the compound is a monoclonal antibody that specifically binds to PSGL-1. m one embodiment, the method includes an additional step of administering an agent that binds te the roonocicnal antibody and induces the cross-linking of a plurality of PSGL* 1 an n g; as on the surface of the T eel], In one embodiment, the method includes inducing the cross-linking of a plu-rality of PSGL-1 antigens on tJis surface of the T cell, wherein the cross-linking indices the signal transduction pathway that results in the death of the T cell. In one example, the method includes a stsp of selecting an individual diagnosed a$ having an autoimmune disease. In another example, the method includes a step of selwing an individual that has received or is expected to receive an allogeneic or xenogeneic transplant. In another example, the method includes a step of selecting an individual diagnosed as having an allergic disease. In another example, the method incli.ctes a step of selecting zn individual diagnosed as having a T cell cancer In one embodiment., the T celi is an activated T cell In one example, the T cell is ■; CIH+ T cell, In another example, the T cell is a CD8+ T cell. In one embodiment, the method includes EI step of detecting the number of T ctffc in a first biological sample taken from the individual before the administration of tbft compound and comparing the results with the number of T cells in a second biclogical sample taken from the individual after the administration of .tie compound. In another embodiment, the method includes a step of detecting a biological activity of T cells fa a first biological sample taken from the individual before the acbiinistraiion of the compound and comparing the results wjth the biological activity of T cslls in a second biological sample taken from the individual after the administration ox the compound. In one embodiment, the administration result in the depletion of at least 20% of p::upherai blood CD3-r cells in the individual In some embodiment*;, tiie adnxistration results in the depletion of at least 30%, 40%, 50%, or n:ore of the peripheral blood CD3+ celU in :he individual. In one embodiment, the antibody oi antigen binding fragment thereof induces I tnt death cf at least 20% o~ peripheral blood CD3i- cells in fhe ijid/vidcal after e>.:;o?;ute to the antibody o.r antigen binding fragment thereof. In some embodiments, the administration induces the dead; of at least 30%, 40%, 50%, or mcie of the pmpheral blood CD3+ cells in the individual Ceil death can be measured at any time, e.g„ one, two, three, four, five, si;c, seven .orxaore days* after exposure to the antibody or £.iidgftn binding fragment thereof. In another aspect, the invention features a method of inducing ths death of a T ceJ; or a natural killer (NIC) cell. The method includes the steps of: providing &T cell or Kl cell expressing PSGL-1 on its cell surface; and contacting the T cell or NIC cell with a The compound used in such a method can include an antibody or antigen bitiving fragment thereof that specifically binds to PSGL-1. In one example, the compound is a monoclonal antibody that -specifically binds to PSGL-1. In one embodiment, the method includes a step of contacting the monoclonal sjitibcdy with an agE-Tit that binds to the monoclonal antibody and induces the cross-linking of a plurality or* ?SGL-1 antigens on the surface of the T cell or MX cell. In one embodiment, the method includes u step of inducing the cross-linking of a pi' jyjJity of PSGL-1 antigens on the surface of the T cell or NIC cell, wherein the cro;:5 0.;infctnc induces the signal transduction pathway that results in the death of the-T c:JIoiNKceIl. In one embodiment, the T cell is an activated T cell. In one example, the T cell hi a CD4-f- T cell, In another example, the T celt is a CDS+ T cell. In one embodiment, the method includes a step of assessing tie viability of the. T VA'A orNK cell after the contacting with the compound. Tn one embodiment, the method includes a step of assessing &. biological activity of ito T cell or NK cell after the; contacting with the compound. In another aspect, the invention features a method of screening for a modulator :>[ ?SOL-l funciion. The method includes the steps of: providing a call expressing RvoL-l on the aurfese of the, cell; contacting the cell with a test substance; and inc curing the viability of the cell after contacting the cell v/ith the t«s: substance to ;'.'•«. rsby determhe if rhe test substance is a modulator of PSGL-1 function. b\ one embodiment, the method include-a the step of detecting ;he death of the :;;;'(induced by the test substance to thereby determine that the test substance is a u.;ckilator of PSGL-1 function. 3:,i one embodiment, the test substance is an antibody or antigen binding fra^iient thereof that specifically binds to PSGL-1. In one example, the test substance is a monoclonal antibody that specifically binds to PSGL-1. In on* embodiment, the me!:ho;l includes the step of contacting the monoclonal antibody with an agent that bine!.. u> the monoclonal antibody and induces the cross-linking of a plurality of PSGL-1 artigens on the surface of,the cell. In one embodiment, the method includes the step of inducing the cross-linking of .t plurality of PSGL-I antigens on the sutface of the ceil, wherein the cross-linking inirii-Kifio the signal transduction pathway that reside in the death of ths* cell. In one embodiment, the T cell is aa activated ll cell. In one example, the T cell is::; CD4+T cell, In another example, the T cell U z, CD8+ T cell. in one embodiment,, the method includes the step of manufacturing bulk qvjkintyiies of the test substance and formulating the vest substance to a pharmaceutical! y &<::cpti:ible-carrier.> In another aspect, the invention features a kit containing: a compound that binds to PS C*Lri on rhe surface of i. T cell, wherein did binding of the compound to PSGL-i OTL the surface of the T cell induces a signal transduction pathway that results in the d:vi:h of the T call; and instructions for use of die compound to treat .autoimmunity, transplant rejection, an allergic condition, or a T call cancer, bi other embodiments, the iii .Tidud-ss instructions for use to treat any disease or disorder descried herein.. An advantage of the invention is that it allow for the depletion of T cells and/or t;:to induction of apopto?is in T cells without causing an associated unwanted or harmful itimu'.ne response. For example, the administrator: cf an snti-PSGL-1 antibody to an individual does not result in an unwanted elevation in the levels of inflammatory cy;o*ines such as .CL-2 or 1MF-& Another advantage of die invention is that it allows for the targeting of a cell surface protein, P^GL-1, whose expression is largely limited to leukocytes, and in p;i';.ti.oiiar T cells Unless otherwise defined, all technical and scientific terms used herein have the ;;an:ci -waning as coirmionly understood by those of ordinary skill in the ait to which this .invention belongs. Although methods and materials similar or equivalent to those? deserted herein can be used In the practice or testing of the invention, soitable methods and materials are described below, All publications, patent applications, patents, and other references mentioned herein are incorporated by reference in their entirety. In c&se of a conflict: in toxminology, the present specification will control, In adiifton, the described materials and methods axe illustrative only and are not intended to ;M? limiting. 'Other features and advantages of the invention, will be apparent from the following detailed description and the claims. Brief Description of the Drawinga ?ig. 1 depicts the results of i time-course experiment that investigated when aeratedT cells acquire sensitivity toTAB4 (an anti-PSGL-1 monoclonal antibody)-mediated apoptotic signals. Fig. 2 deplete die results of cell surface bioiiriyiation and immunoprecipitation of : r;>ii antigen recognized by the TAB4 antibody. Fig. 3 depicts the expression of thePSCL-1 airigen on spleen CD44-T cells, CU8+ T cells, CD19+ B cells, and NK cells. Fig. 4 depicts the expression of tlie PSGL-1 antigen on CD4*S CD8\ and CJ>l+:r, and C3XT thymocytes. Fig. 5 depicts the levels of IL-2 produced in mixed lymphocyte :ulture asing spliiMTt cells isolated, from TAB4 (or hamster Ig)-tfeated Baib/c mice as tl*ie responders and clli.-rn3sniatched C3H spleen cells as the stimulator. Fig, 6 depicts western blot analyses demonstrating that (A) proteins in ji.iuLOpjcecipitatsd with the TAB4 antibody can be recognized by a commercially av;;:J;jl)le anti-PSGb-1 antibody and (B) prehearing of T cell lysate wth anti-PSGL-1 antiboiy can deplete the proteins recognized by the TAB4. Fig. 7 depicts the percentage of surviving grafts in C57BI./6 rai-a that received a :is:in graft* from Balb/c ini::e and were treated wirh an anti-PSGL-I antibody (closed d:;:'r:i:);;id) or a control antibody (open square). Fig. 3 depicts the time course of the percentage of apopiolic T cells following the ticafcnent of activated human peripheral blood nononuclear cells w;.th an auti-kanMii'i T'SGL-1 antibody. Fig, 9 depicts the incidence of diabetes in antoitomune non-obese diabetic (NOI.1) male mice that were treated with anti-PSGL-1 antibody (closed square) or a cor^o] jntibody (open square), Detailed Description The invention is directed to methods of modulating T cell activity by me:! ilAdnjg the function of PSGL-1 molecules residing on the surface of a T cell. Engagement of PSGL-1 with compositions described herein can cause the depletion of T cfil'M and'or induce T ceils *:o undergo apoptosis. These compositions arc ihercfoie useful as therapeutic agents for controlling immune-related conditions such as autoimmune diseases, transplant rejection, allergic diseases, and/or T cell-derived ca'osarc. The compositions arcs also useful in causing the depletion of T cells from acy biological sample where the presence ox activity of T cells is not desired. PSGL-1 is ? cell surface adhesion moleculs ".hat is expressed on neutrophils, T ?.) id B-lymphcsiytes, NE crils, monocytes, dendritic cells, and primitive human Ct>34 hematopoietic progenitor cells. Through its ability to jmeract with seleatins, PSGL-1 mediates the rolling of leukocytes on the endothelium and the extravasation of lev.kocyifi5 into raff.amed tissues PSGL-l-rnediated binding of T cells to E- and P-sek-cva, or migration, is differentially regulated. For instance, the. appearance of CLA (cupreous .lymphocyte ant gen) epitope is thought to be induced on T cells undergoing naive to memory transition. Only activated helper 1 but not helper' 2 T cells express functional PSGL-1 and it capable of migration into die inflamed area of the skin. PSGL-1 is a sialomucin that must he specifically sialylated, fucosylated, and binding activity appears to be s. result cf differential past-transialionai modification, as suggiustsd by elevated levels of alpha (1,3) fucosylii&nsferases activities in activated T celk- P'SGLrl isofonns also show differential affinity to L-setectin and E-selectin. For msun:.cw, human T cells exhibiting the CLA-positive isoforai can tether and roll on both B- iind. P-selectin, while T eel la expressing PSGL-I \vi:hout the CLA epitope only bind to .P-i-eJsotiri. Furthermore, binding of PSGL-1 tc P-selectin is contingent upon the piesence of the terminal decapeptide that contains threa tyrosine residues for sulfadon ana OIK: threonine residue for glycosylation, A PSGL-1 protein can be prepared by recombinant methods and/or by isolating a TiFt.i-ve PSGL-1 protein from biological material A recombinant PSGL-1 protein can be ]i::c:iuced in prokaryodc or eukaryotic tells, either in vitro or in vivo* Nucleic acids ens:: ding PSGL-1 can be used for recombinant production of the protein (see, e.g., GmUankO Accession NMJ303006 for an example of a nucleic acid deeding a PSGL-1 polypeptide). Antibodies directed to PSGL-1 are also well known and can be used for purification of the antigen (see, e.g., Herron et aL (2000) Science Juo 2;i:" 3(;:i47l); 1653-56; WO 0Q'25B0S) and/or used in Methods described herein. PSGL-1 if' /uither described in references including but nor limited to Sako et al, (1993) Cell 75:117?; Vachino et ah (1995) J. Biol. Chan. 270:21966; and Vekkn ;n ec d, (1993) J. B.::i.:::rieml 270:16470. For recombinant production of PSGL-1, the siamltsaeous expression of both P': CL-1 and its modifying alpha- (1,3) fucosylb-aMerase, Fuc-TVU, umy be required t\v ih» functional expression of FSGL-1. In addition or alternatively, recombinant pr:.c.cMion of PSGL-1 may be accompanied by co-traasfection with a nucleic acid encoding PACE for removing the propeptide and/or or a nucleic acid encoding tyrosine sulfoUansferase. An anti-PSGL-1 antibody can be used to isolate and purify a PSGL-1 antigen flora ecological na'cerial. A.:ay cell type expressing a PSGL-1 protein, e.g., X cells dc.iiv-ir-ii from an individual or a T cell line, can be used as a source of the protein. Once p.died, the protein can be used m a variety of methods as described herein. For e>':urrple, the purified PSGL-]. protein cars be used in an in vitro screen of moaulatots of PSCL-l function 0"?. T cells or as an iiamunogen to prepare antibodies directed against In addition aPSGL-1 antigen can be purified using a selectin-Fc fusion, e.g., a P-s e!::et:n-Fc fusion. Ask. PSGL JLA mibodies. PSGL-1 polypeptides (or immunogenic tfragatsnts or analogs thereof) can be use:;, io generate antibodies useful in the methods of the invention. As described above, PSGL-1 polypeptides or peptide fragments thereof cin bss produced by recombinant teeluixj;.es or synthesized using solid phase synthesis methods. The recombinant PS0L4 polypeptides or a peptide-; fragrant thereof can be used. a3 an iirmunogen to prod.xc: anti-PSGL-1 antibodies. In addition, an anti-PSGLrl antibody, such as the TAH1 monoclonal antibody, can be used to purify a PSGL-1 polypeptide, e.g., a PSGL-1 pvlypsiptide in its natural conformation, which een then be used as an immunogen to prtfdi:-c* additional anti-PSGLrl antibodies. An antibody of the invention can be a monoclonal polyclonal, o:: engineered anybody that specifically binds to a PSGL-1 polypeptide. An antibody that !'spE.dfi::aily binds;1' to a particular antigen, e.g., a PSGL-1 polypeptide, will not subii;;i;v:ially recognize or bind to other molecules in a sample. Thus, the inventior also fei;l;.i:cj: methods tor identifying a test compound (e.g., an antibody) thst binds to a polyjiq-tide of the invention by contacting Che polypeptide with a tesr compound and de;n::T:inmg whether the polypeptide bind* to che test compound (e.g., by direct deca, fion of the binding, detection of a competitor molecule- which disrupts binding of the i-si compound to tlie polypeptide, and/or detection of binding using an assay for apoptcsiis-indudns activity). In general, PSGL-1 polypeptides can be coupled to a carrier protein, su'ch as KL.r*. rixed with ar. adjuvant, and injected into a host mrarmal. Antibodies produced in ■■Jv.d: minimal can then be purified by peptide antigen affinity chromatography. In particular, various host animals can be immunized by inject on with a PSGL-1 f ):ypeptide or an antigenic fragment thereof. Commonly employed host animals inca-ii i&bbits, mice, guinea pigs, and rat1?.. Various adjuvants tha ca;i be used to inert £3;:i the immunological ■response depend on the best species and include Freund's adju /:?nt (complete and incomplete}, roineral gels such as aluminum hydroxide, surface active irobstances sush as lysolecithin, phrenic poiyols, polyar/ions, peptides, oil eir.x *ions> keyhole limpet htsanocyanin, and dirut:ophenol. Potentially useful human adjuvants include BCG (bacijle Caknctte-Ouerin) and Coiynebacteriuin pawum. Polyclonal antibodies are heterogeneous populations of antibody molecules that are coririn.sd in the sera of the- immunized animals, Antibodies within the invention therefore include polyclonal antibodies and, in a Monoclonal antibodies, which are homogeneous populations of antibodies to a patii..;uiar antigen, can be prepared using thePSGL»l f olypeptides described above and st&r.'L&rd hybridoma technology (see, for example, Kohler ex al, Nature 256:495 U^'ST. Kohler et al., Eur J Immunol 6:511 [1976]; Kohkr et a.L, Eur J Immunol 6:292 Ilfi/'o], Hainmeiiing et al., Monoclonal Antibodies and T Cell Hybridamas.. Elsevier, NT. | :9S5J). In particular, monoclonal antibodies can be obtained by any technique that prov idj-s for the production of antibody molecules by continuous cell .Sines in culture such a; described in Kohler ist al, Nature 256:495 (1975), and U.S. Pat. No- 4,376,110; the :.:nman B-cell hybridoma technique (Kosbor si: al./Immunology Tsday 4:12 [1983]; Co,:: Once produced, polyclonal or monoclonal antibodies are tested for specific Pi>GL-l recognition by Westwrn blot orimmunoprecipiiation analysis t»y standard mg-tiiods, for example, as described in Ausube] et al., supra. Antiboftea tbat sp-.-.,j:fical!y iecojjniie and bind :o PSGL-i are useful h die invention. Afiti-PSGL-l ai\'ibcdieEi that bind to the PSGL-1 antigen on the surface of a T cell, e.g., a CD3+ cell, an-i induce the depletion and/or apoptosis of T cells b an individual firs particularly uk\nJ. The antibodies can be used, for example, as part of a therapeutic regime (e.g., ic rfiduoe o: eliminate an undesirable immune response, such as a T cell mediated inuauns jx'spcmse, associated with conditions such 34 autoimmune- diseases, transplant rejection, allergic diseases, and T cell-derived cancers). Antibodies also can be used in a screwing assay to measure the. ability of a candidate compound to biud to PSGL-1. Li addition, technique;; developed for the production of "chimeric antibodies" (Monison et ai: Proe Natl Acad Sci USA 81:6851 [1984]; Neuberger et ai,, Nature 312:604 [1984]; Takeda et al., Nature 314:452 [19845) by splicing the genes from a moiiS'ii antibody molecule of appropriate antigen specificity together with genes from a hum an antibody molecule of appropriate biologies! activity can be used. A chimeric antibody ia a molecule in wh::ch different portions are deiived from different animal .spe-d'-ss, such as those having a variable region derived from a rnurine raonoclonal antibody and a human inmuttoglobulin constant region. Alternatively, techniques described for the production of single chain antibodies (US. Put. Nos. 4,94-5,778,4,946,778, and 4,704,692) can be adapted to oroduce single chs-ir antibodies against a PSGL-1 polypeptide, or a fragment thereof. Single chain antibodies are formed by linking the heavy and light chain fragments of the Fv region via a. a amino acid bridge, resulting in a single chain -polypeptide. Antibody fragments that recognize and bind 4;o specific epitopes can be generated by known techniques. For example, such fragments include but are not Hnu'ici-. to F(ab')2 fragments that can be produced by pepsin digestion of the antibody tno"i':!Cule, and Fab fragments, tha1; can be generated by reducing the disulfide bridges of F(;ib )2 fragment*;. Alternatively, Fab expression libr,siies can be constructed (Huso ?;i ai., -Science 246:1275 [198911) to allow rapid and easy identification of monoclonal Fah fr«;2 -rui-nts with the desired ipeciScity. Antibodies can be humanized by methods known in the on. for example, ir :nodonal antibodies with a desired binding specificity ran be commercially htvriiiikized (Scoigene, Scotland; Oxford Molecular, Palo Alto* Calif. \ Fully human aiuihodies, such as those expressed in transgenic animals are also features of the invention (Green et al> Nature Genetics 7:13 [1994]; and U.S. Put. I Sordine;iAssaysifor Compounds that Modulate PSGL-1 Function Tiie hi vent: on also encompasses methods for identifying corr.pounds that h'r;i:sM':3t with PSGL-1 (or a domain ofPSGL-I) including, but not linked to, cenpounds that induce T cell dspktron and/or T cell apoptosis upon binding to PSGL- \, Aho included are compounds that modulate the interaction of PSGL-l with transmembrane, extracellular, or intracellular protons that regulate PSGL-l activity and ::>[i;.pounds which modulate PSGL-l activity. The compounds that may be screened in accordanee with the invention include, but ;ii'c; :::.ot limited to peptides., antibodies and fragments thereof, and other organic coiMpci i.nds that bind to PSGL-1 and modulate a biological function mediated by PSGL-l, as described herein. Such compounds may include, but are not limited to, peptides such as, for exwj'ipto, soluble peptides, including but not limited -.0 members of random peptide liberie:-; (Lam et al„ Nature 354:82 [1991]; Houghton et al„ Nature 3:54:84 [1991]), and combinatorial chemistry-derived molecular library made of D- and/or L corf:juration amine acids, phosphopeptides (including, but not limited ':o, members of rand ;>:«:. or partially degenerate;, directed phosphopeptide libraries; Songyang et ai., Cell 7£:767.[19933), antibodies (including, but not limited 1.0, polyclonal, monoclonal, huiniuized, anti-iiiotypic, chimeric or iingle chain antibodies, undFAb, F(ab')2 and FAb expression .library fragments, and epHope-bixiding fragment? Thereof), and small or^fii'aic or inorganic molecules. Other compounds which can be screened in accordance with trie invention include: but art; not amked to small organic molecules that affect an activity of the Pill II ,'1 p.rote.rn, as described herein. Computer modeling and searching technologies permit identification of compounds, or ihe improvement of already identified compounds, that can modulate PSCiL 1 expression or activity. Having identified such a compound or composition, the adi v •; sites or regions are identified. Such active shes might typically be a binding site fo* a natural modulator of activity. The active sit;? can be identified using methods known in the art including* for example, from the asirao acid sequences of peptides, fzcw the nucleotide sequences of nucleic acids, or from study of complexes of the r:li:-v3.nt compound or composition with its natur.il ligand. In the latte:: case, chemical or X-ray crystallography methods can be used tc find the active eite by finding where or. i)w: factor the modulator (or ligard) is found, Although describee, above with reference to design and generation of compounds which could alter binding, one could also screen libraries of known compounds, including natural products or synthetic chemicals, and biologically active roat(! :i als, including proteins, for compounds which bind to a PSGL-1 protein and cause T cell depletion and/or indues T ceil apoptosis, h vitro systems may be designed to identify compounds capable of interacting with PSGL-1 (or a domain of PSGL-1), Compounds identified may be useful for exa^ioit', in modulating T eel! activity as described herein and thus may be useful for \he ttofl-tment of conditions such as autoimmune diseases, transplant rejection, allergic c!isc;;i:(;i;r andT cell-derived careers., The principle of the assays used to identify compounds that bind to PSGL-l invr-ivca preparing a reaction irtixture of PSGL-1 (or a domain thereof) and the test compound under conditions and for a time sufficient to allow the two components to intec -id fjici bind, thus forming a complex which can b« removed and/or detected in the rem;uou mixture. The PSGL-1 species used cat) vary depending upon ths goal of the scTCif-sning assay. In some situations it is preferable to employ a peptide corresponding to a domain of PSGL-1 fused to a heterologous protein or polypeptide that affords adwvn.tiigeE; in the assay system (e.g., labeling, isolation of die resulting complex, etc.) C*\J ':>■:> utilized. Ths screening assays can be conducted in a variety of ways. For example, one rri-t 101 to conduct such an assay involves anchoring PSGL-1 protein, polypeptide, p.r:)' Lb or fusion protein or ths test substance onto a solid pha.se and detecting PSGL-1 /t;j r, compound complexes anchored on the solid phase at the end of the reaction- In on: embodiment of such a method, the PSGL-1 reacuim may be anchored onto a solid s^d'p.C':?, and the us; compound, which is not anchored, may be labeled, either directly or i Erectly, "In practice, microtitsr plates may conveniently be utilized as the solid phase. Tlr anchored component may be immobilized by non-covalsnt or covalent &.::i:ich:coents. Non-covalent attachment may be accomplished by simply coating the s;:!!(! iiurfaoewith a solution of the piotein and drying, Alternatively, an immobdized ;m' Lbody, preferably a monoclonal antibody, specific for the protein to be immobilized r:ii:.) :;ie used to anchor the protein to the solid surface. The surfaces may be prepared in jjd /;;.ji.ce and stored. In order to conduct the assay, the nomminobilized component is added to the ■;:c :r:t:d surface containing the anchored component. After ths reaction is complete, ursveacted cornporente are removed (e.g., by washing) under conditions such that any c:or minxes formed will remain immobilized on ;he solid surface. The aer.eetion of conpb: Alternatively, a reaction can be conducted in a liquid phase, the reaction produce separated from unrsactsd components, and complexes detected; e.g., using an imrr:civilized antibody specific for PSGL-1 proton, polypeptide, peptide: or fusion prot'Mr. or the test compound t3 anchor any complexes formed in solution, and a labeled anybody specific for the other component of the possible complex to ietect anchored co['ftp>!xe&. Alternatively, cell-biased assays can be used to identify compounds that interact with PSGL-i. To this end, cell lines that express PS&L-l, or cell lines that have been genetically engineered to express PSGL-1 can be used, Cell based assays are pa:lioUarly useful for evaluating the functional effect of a compound identified by a sccean described hssrein, Fo:; oxample, once a compound is identified based upon its al: ilJt;y to bind to a PSGL-1 protein, the compound can then be tested fov it? ability to, c. ;!'... induce T ceil upoptosh; in vitro or in vivo or deplete T ceils in vtro or in vivo. Pharmaceutical CopjgositicQs, Given that an objeciof the present invention is ro alter an immune response in an individual, a pharmaceutical composition containing, for example, antibodies, small r^olecules, or othe:: compounds that specifically bind PSGL-1 polypeptides aie also a fe:ij"i;ui-3 of the invention, t.. a preferred example, the compound functions as an agonist ofPSGL-l. Pharmaceutical compositions for use in accordance with the prssant invention car. o? formulated in a conventional manner using ei:.e or moie physiologically a:cep:ablB canters or excipjente. Thus, the ccrnpoiinis and their physiologically a:<: enable salts and solvates may be formulated for administration by a variety of routes admimstratfon.> The compounds may b& formulated for parenteral administration by injection, for 'Sxanipte, by bolus injection or continuous infusion. Formulations for injection may be plaited in unit dosage form, for example, m ampoul&s or in multvdose containers, wji:h as added preservative. The compositions may take such forms a fyfefhods of Copb-o.;3i>ig a T. Call-Mediated Immune Response and Depleting T Cell l!2Eyr,E23Q£ Compound:; such as those detailed in the screening* assays described herein may be xi&eful, for example, in modulating a biological function mediated by a PSGL-i poly/spride and/or for the treatment of disorders associated an excessive or unwanted immuns response, e,g., a T cell -mediated immune response. These compounds include, but a:'« not iitxrited to peptides, antibodies and fragments thereof, and other organic compounds that hind to PSGL-1 en the surface of a T cell and induce a signal trar.!;::iu'.;tiOM pathway that results in the death of tins T cell. The methods of the uivin: lion optionally include the addition 01 a cross-linking agent that induces the cross-lirdti'ii-g of PSGL-1 on the surface of a cell. The compounds described herein can be used in my instance wherein the depletion or clixamnrion of T cell activity is desired. Pani-::ul.arly useful condition;; that can be treated with the compounds of the invention i»cltrd.'j autoimmune, diseases, transplant rejection, allergic diseases, and T cell-derived Examples of condition:; that can be treated with the anti-PSGL-1 compounds described heroin include, ha: are not limited to, diabetes msllitus, arthritis (including rh:::!i:.Mtoi(i arthritis, juvenile rheumatoid arthiiust osteoarthritis* and. psoriatic arthritis), mtiliipie sclerosis, encephalomyelitis, myasthenia gravis, systemic lupus erythematosa, au;oijx.mune thyroiditis^ dermatitis (including atopic dermatitis and eczematous dfirmanria), psoriasis, SJLJg^ns Syndrome, Crohn's disease, aphthous uker, iritis, oortiaiiotmiis, keraaconjunefcvitis, type I diabetes, inflammatory bov-zel diseases, ulccraive colitis, asthma, allergic asthma, cutaneous lupus erythematosus, scleroderma va^:ni;isa proctitis, drug eruptions, leprosy reversal reactions, erythema nodosum le;;.iosum, autoimmune uveitis, allergic encephalomyelitis, acute teciotizing hemorrhagic encephalopathy, idiopathic bilaieial progressive sensorineural hearing lof-E, E.plaistic anemia, pure red cell anemia, idiopathic thrombocytopenia, polychondritis. Wegener's granulomatosis, chronic active hepatitis, St&vens-Johnson. syndrome, idiopathic sprue, lichen planus, Graves' disease, sarcoidosis, primary hi Is ar chihesis. uveitis pDsierior, interstitial lung fibrosis, graft-versus-host disease, casts of t.?: implantation (including traasplantation usir-g allogeneic or xenogeneic tissues) such a.>; '::>o:\c marrow transplantation, liver transplantation, or the transplantation of any organ or tissue, allergies such as atopic allergy, AE>S, and T-cell neoplasms such as J?ul:£iini.i:is and/or lymphomas. The methods of the invention can be used to deplete T cells firoici a cell popul ation, either in vitro or in vivo. For example, a biological sample derived from an individual can be depleted of! cells in vitro by contacting the sample with an anri-PSttL-1 compound described hsrein, optionally together with a cross-linking agent Thin :r;ie:hod can bs useful, e,g.f by allowing for the enrichment of non-1' cells in a ceil popixl.ii.i on as well as by reducing or eliminating T cell activity from a cell population. The following are examples of the practice of the invention. They are not to be construed as limiting the scope of the invention in any way. Bxftmplfcs Examine 1.:. Preparation of an Anti-T Cell Apoptosja Inducing ffrctein rTAIP!!) MvJlclonal Antibody A TAlP-specific wortodonal antibody was. generated by applying the well fcnov.'n eel] fusion methods of Kohier and Milstein ((1976) "European Tournal of bxur.iunolo.gy 6:511-519) to produce a hybridoma secreting desired antibodies. Aj'tjbo'ly producing cells from a hamster injected with Concanovalin A (Con A)» acl:i\ &bii Balb/c tpleen T cells were fused with a aiycloiua celi line to form an anrihuJy sec:r:-;ii:;g hybridoma. The two populations of cells were fused with polyethylene glycol, and the resulting antibody producing cells were cloned and propagated by standard tissue culture methods. One hybridoma generated according to theso methods S6:rxcd a monoclonal antibody, designated TAJ34, thai was able to induce T cell apootasis in vitro and deplete T cells in vivo. The protein recognized -?y TAB4 was designated T cell apopiosis inducing protein (.TAD?). C57BL/6J (B6) and B ALB/c mice were purchased from the Jackson lab (Bar Hrtfbor, ME). Syrian hamsters were purchased from the Animal Core facility, National liu-.v3.ft University Medical College. Concentrated culture supernatant of the TAB4 hybridoma was spun at 20,000 x g for 10 minutes and the supernatant was diluted at a 1:1 ratio with the binding buffer (0 I M sodium acetate, pH 5,0). A protein G column (approximately 1 ml bed volume) was, wished three times with 3-5 ml of the binding buffer. The cleared culture supcs ns.tant was loaded to the protein G column and the flow-through wus collected and rdoadsd to the column. The column was washed with 6-10 mi of the bidding buffer anci the bound antibody was dated from the column with 5 mi of the eiution buffer (0,1 M ,1'lysi.ne-HCl, pH 2.3). Each fraction contained 1 m] of the eluted antibody and the elulsd traction was adjusted to neutral pH by mixing each 1 ml fraction with 50 microliters of 1 M Tria-HCl.. pH 7.5. Fractions containing the antibody ware pooled and dia ■> zed against 2 liters of FB.S, pH 7.4 three limes at three hours for each dialysis, Protnir. concentration in the antibody samples were determined with the procedure devilled by Bradford using tlie Bio-Rad 'Protein Asssy (BIO-RAD, Hercules, CA)~ F:A ;i'C.[ile ?.: Preparation of a Mouse Spleen Cell Suspension and.the Activation and fii:i*:chment of T cells Mouse spleen was immersed in S ml of Hank'.5 balanced salt solution (HBSS)S geaily minced with a sterile cover slip, transferred ta a 15 ml centrifuge tube (Costar), and spun at 200 x g for 5 minutes. The supernatant was discarded and ';he cell pellet w :"f. resus;?ended in the residual buffer by gently tapping the wall. The contaminating re:l bli.>od cells (RBC) were lysed by the addition of 1 ml of RBC lysis buffer (0.6 M NH ;.C1; 0.17 M Ms-base, pH 7,65). followed by a 2 ntiu incubation at room temperature and rapid quenching with 9 rnl of BBSS. The cells \v«e pelleted at 200 x g -jtr 5 minutes, washed twice and resuspended in RPMI medium. Tnt concentration and viability of cells in the mixture were determined with a hemocytometer (Cambridge S:i'in ific Inc.) and Trypan biue exclusion. Tha spleen cells were adjusted to a final concentration of 3 X 10 /mi with KPMI milium and Ccmcanovalin A was added to a final concentration of 2 miorogranis/inl to z.:tlvi\xz the T cells. The cell suspension was transferred to a 6->velI srlturc plate (;:•' Tih'well) or a Id-cm culture dish (10 ml/dish) and incubated at 37X; 5% CO^ for 4S hows before harvesting. The activated spleen cells, including activated T cells, were ■^suspended in 5 rnl of KB .3.8 and carefully overlaid on top of a 5 ml 55% cushion of ?e:xoti solution in a centrifuge tube, Care was taken not to disturb tne separated lay ere. The ctite were spun a). 1,900 x g for 13 minutes at 25CC without the brake. The '■:ii;:it;hed T cells were collected iron the interface of the two layers, washed twice with I'ilJSS, and were teady for experiments* W&Zlpk? 3.;.. Apoptpsis of Activated T cells /activated T cells (see Example 2) were resuspended to a final concentration of 3 X 'Jf cells/mi in RPMI meclirin] containing 5 ng/jnl cf H-2, and treated with control ]g, TAB 4, or anti-CD3 according to the conditions shown in Table 1, After an incubation period of 18-24 hours, the extent of apoptosisi in each alibis v/as determined using the 7-AAD apoptosis assay. The treated cells were trariiifsivsd r.o FACS tubes (Fdcon), washed twice: with ice-cold FACS solution (1% jfete'l bovine serum, 0.05% sodijsn azide in PBS), pelleted at 200 x g at 4CC. The cells vrer;: :ej.^sperided in ice-cold FACS solution re a final concentration of* 1-2 X 107 celhfrd. For staining, 0,1 ml of the resuspendec. ceils were mixed with 7-AAD to a :;inaJ ;:oncentration of 2 ug/ml and then incubated a*; 4°C in the dark for 2.0 minutes. Finally, :he stained cells were washed twice with ice-cold FACS solution, resuspefidad :;n 0-5 :;cl of FACS solution and analyzed with BD LSR flow cytomeier (Beckton 'Dic'rli.fOi'O. Fig. 1 depii;ts. the results of a representative time-course experiment that •investigated when activated T cells acquire sensitivity to TAB4 (anti-TAlP)-mediated apo^totic signals. Mouse splcnocytes were activated with Con-A and nwiiAtsined in IL-1 c raraniittg medium. Activated T cells were harvested, resuspended, and challenged with TAJB4 monoclonal antibody or control hamster IgG in the preserve of anti-hamster !gG anybody as cross-linker. The ability of TA1P ero&s-tinking to induce low level (6.5^) of apoptod; cell death was evident on day one. However, the ?xv.enr of TAB4-induced apoptosis increased ;;rom 17% on day 2. peaked at 52% on day 4S ar.d declined to 44'* on clay (5. The control haraster IgG did net induce specific apotuotic T cell death. a.& compared with the cultures that Teceived only EL-2, Antt-CD3 (as positive control) induced apoptosis in 38% of T cell after 48 hours of activation (data not shon^i). jSxa^pk^JBffiression.oi^ Cells wers washed twice with ice-cold FACS solution (1% fetal bovine serum, 0.0?* sodium azide in PBS) and spun at 200 x g at 45C in a FACS tube (Falcon), The cel!:j ivere restispendad in ice-cold FACS solution to a final concentration of 1 x 10' celled, and a 0.1 ml aliquot of the resuapended cells ir. a FACS tubs (Falcon) M»as us£d for each assay. For surface staining, the TAB4 monoclonal antibody or a control haxfusi a? Ig at a final concentration of 2 u&'ml were added to the cells and the mixtures weri- \nc: abated at 4°C for 30 minutes in the dark. The cells were wasted once with ice-sold 1-ACS and then stained with: (1) for spleen cells. :ychrome-conjugated airti-CD3 antitodv (2 ug/iritX FITC-co:vjugated anti-hamster Ig, and PE-conjugaced and-CD-!iyCIM/CD19/C»llb/pan«NTCl-A/I-E/Mac-3 antibody (2ugfcil) in 100 ul ofice-:*.o.W ?ACS solution; and (2) for thymus cells, FITC-conjugated anti-hamster Ig, PE-conj'j.gfued anli-CDS, andcychrome-conjugated antUCD4 antibodies (2 jg/ml) in 100 ul of JC2"COM FACS solution, Ths ieaction was performed at 4°C for 30 minutes in*he dark. Finally, the stained cells were washed twice with ice-cold FACS solution, resinsiwnded in 1 nil of FACS solution and analyzed with. BD X.SR flow cytooieter (Bed tei DicKhon). Figs. 3 and 4 demonstrate by FACS analysis the distribution ofTAlP antigen en ths .';in.ous subpopulatioa3 of spkuocytes and thyraocytes. As shown ir. Fig 37 CD19' B eds expressed low but detectable amounts of TAIP proteins on the surface. * Sig::i"3«3.!mtly higher amounts of TAIP proteins weie detected on CD3+ T cells and a fra; ii :m of NK ceils. Most of the CD4", CDtf. and CD448+ thymus T cells expressed significant amounts of TAlP proteins. In contrast, ths TAIP proteins were expressed on; y on a small population of CD4T thymus T cells (Fig. 4). 'Tissues ftom B6 and BALB/c mice, including brain, thymus, hesirt, lung, liver, dtonucli, kidney, splssn, and skin, were collected, fixed in 10% formaldehyde evenly as. room temperature, and embedded ir. paraffin blocfcs. Tissue sections, at a 4 uni trictoess, w spread in 45 9C water, and laid on a coated slide. The slides were dried in T7°C and were reedy for subsequent experiments. Slides containing the tissue paraffin sections were dewaxed and dried through a xyl-Mi ;JH- 100% eths.no} series according to standard protocol and were finally kept in I00:;J: eihanoh The sections were rehydrated through a 100% ethaacI-90% ethanol-85& vrtfcatiol—70% ethanol-PBS serial incubation according to standard protocol to a final PBS solution. The following reactions were til performed in a humidified box. Nonspecific binding v/ere. blocked oy incubating the tissue sections in Mocking buffer (1% r.oimal goat serum) for 1 hour at room temperature (or 4"C overnight). The bloebin;;>; buffer was removed and TAB4 or normal hamster Ig (1:200 dilution) was added to the? sections and incubation continued for another hour at room temperature (or The results indicated that the TAB? proteins expression were detected only in 3on-5 niiarcw derived-tissues, but not on the rest of the tissues tested Exsrop'-ieS.; Call Surface Biotmvlation and lTnrauno3recipitauon_Qf .the TAIP Autiaen 5 x 107 RL Labeled calk were lysed at a density of 5.C x 107 cells/ml in cold lysis buffer '1% Tr.ton X-10C, 20 mM Tris-HCI, pH £.0,160 mM NaCl, 1 mM C;iCl2) containing :ci-iV,y..M.e protease inhibitor c-oclctaii (Roche) for 15 minutes, and insoluble raateiial was jeilcPMi at 10,000 X g for 10 minutes; these and all subsequent steps were performed at 4°C, For immuaoprecipitation, thelysate was pjeiricnbatedfor 30 niin-ites with 50 //J of packed protein G-S spharose (Amersham Pharmacia Biotech) to remove noit-speriilcally binding proteins. Beads were pelleted, and aliquots of the supernatant (ro«nsly corresponding to 5.0 x 107 cells) were incubated with 20 a] oJ protein (i» Seph:iicse prsloaded with 10 (ug of mAb TAB4 or IgG from norma! hamster serum. After hcubiition for 4 h at 4 WC, the resin was washed four times with washing buffer {Q.W% Triton X-10C, 50 mMTris-HCl, pH 3,5, 400 mMNaCh 1 inM CaCl2, J nig/ml ovf;f;»amin)t twice vtfth a similar washing buffer, containing 250 mMirw:ead of 400 ivM Ne.Cl. Proteins specifically bound to the TAB4- were eluted with 50 ^1 of IxSrS ■■ample buffer. Eluted proteins wer-s separated by 8 % SDS-PACE and ttamcer/ed w nitrocellulose membrane (Millipore), Filters were analysed for bioiir ybte.d proteins with peiCEidase-conjugated Avidia (PharMingen) and developed wit!* iks Cheixiiiuminescence reagent (MEN1*"1 Life Science Products). As shoxvn in Fig. 2, a biotinylated surface protean with a molecular weight of approximately 120-kD was identined by TAB4 m Rl£31 cells (TAIP* T cells), but not in XB cells (TA1F" cells). In contrast, protein G sephatwe coated with hamster normal aeru-ji could not irtrieve this 120-kDa proxein. These results suggest that this 120-3 .Exg.rr.pte 6: Depletion of T Ceils in vivo To examins-the effect* of TAB4 on populations of T cells and other cells in vivcK iricc were injected with 300 ug of TAB4 or control hamster Ig intraperitoneal!) and, <: day splenocytos thymocytes and peripheral blood mononuclear cells were harvested for the total cell count exid fox analyses of csll surface markers by facs.> ?or FACS assays, the crfls were fixed with 2% paraformaldehyde at 4°C for 20 J:.V:elites, washed twice, and resuspended in ice-cold FACS solution to a final cottctfT.fcatiott of 1 K IQ7 cells/nil. A100 ul aliquot of the T&suspended cells in a FACS cube i f alccnV) wa^ used fox cash assay. TAB4 or control hamster Is at a final coiif^rtcarion of 2 u&'ml war;:; added to the cells and the mixtures were hviubated at 4*C for 30 minutes in the dailc The cells weie washed once witii ice-cold FACS and teamed with: (I) for spleen cslte, cychrotne-ccnjugated aati-CD3 antibody (2 ug/ml), FIIC-conjugated fnii-hamster Ig andPE-conjugated smti'CD8/CD4/CD19/CDilb/pan- NK/I-Ajl-EyMac-3 antibody (2 ug/ml) in 100 ul of ice-cold FACS solution; and (2) for thynus cells, HTC-conjugated anti-hamster Igs PE-conjugated anti-CD 8, and cychimne-conjugsted anti-OX antibodies (2 ug/ml) in 100 ul of ice-cold FACS sol'jt icii. The reaction was perfomiec at 4°C for 30 manures in the dark:. Finally, the stainM cells were washed twice with ice-cold FACS solution, resuspe:ided in 1,000 ul of !r.\CS solution and analyzed with BD LSR flow cytometer (Beckton Dickison). Four days after the infection, the percentages of CD34 T cells in peripheral Wood leukocytes (P3L) decie&sed from 36.7% m control mice to 4.1% in TAB4-txo?.l y\ mice (Table 2). TAB 4 treatment caused a slight reduction in the total number of sp]«i:. .BX!jr;.ii:i'.;e 7: Anti-TAIP Antibcdv dogs not Induce JLr2 or TNF-q Secretion Balb/c mice (H-2d) were intraperitoneally injected with 300 micrograms of TAB4 or control hamster Ig. Splenocytes wete isolated 7 days after injection, and. used as i:ij:;ponders in culture wih mitomycin C-treated C3H (H-2k) splcnosyles (as .stimulators)* Three day& late,:, the culture superna^ants were harvested md the IL-2 coiu::iit was measured by EOS A set (PharMingenh A? shown in Fig. 5, the IL-2 prodicdon was suppressed in respouder cells derived from TAB4-tr$ated mice as cotiipsu'ttd with that of control mice. The plasma levds of IL-2 and TNF-a were also 2iiiL'yzed and no significant difference was noted in flics levels of IL-2 fcr TNF-a) in tli« sestf-. :)ftiie control and the T.-Ji4 treated "mice. Sh:ce production of IL-2 is central to ilia acta vity of T cells, the resits show that a TAIP-specific antibody, such as TAB4, ca-ci be used i?* v/v,? to manipulate T cells and control unwanted T eell-rr.edip.tod immune responses such as thoss associated with autoimmune diseases arid txan;!].'l:tatatio^ rejection. Bxairi;c.[;::.3_: Usa_oj an Anu-TAJP Antibody to Prevent Transplant Rejection .Vlice (obtained from Jackson Laboratory) at 8 to 12 weeks of a^e were taetfthotiized with Acepromazin indite (Fennenta Animal Health Co., Kansas City, MO'. Frio; to skin grafting, Ticn-ihymectomized recipisnt C57BL/6 mica (H-2b) were injected intraperitoneal^ with :500 ug of TAB4 or isotype control antibodies seven days 'uef 00 skin transplant smgery. Severn days later, a lateral flank of skin from fully aIk)j:*-iMi.«:dc mismatched Balb/cj rales (H-2d) was graftsd. on the lateral flank of the antibody pre-tieated C57BL/6 mice. Seven days post transplantation, the mice were agtb. fc;,ected with 500 ug of TAB4 or isotype control antibody. The mice were tnc: ji'ored every day after graft transplantation. The grafts "were considered rejected \vheri 50% donor skin was necrotic. The percent of graft survival is shown in Fig. 7 (n-sS;. The data show that TAB4 antibody treatments prolonged the survival of the alk;}i:A!'3ic skin graft-3. £xtr::i.ple 9' Identification of TA1P as PSGL-I P-selectin glycoprotein ligand-1 (PSGL-1), also named CD 162, is the main P-siE].:3i:;:;in ligand expressed o:n leukocytes, including T cells (Sako et al. (1993) Cell 751 H9; Vachino et ah (1995) J. Biol. Cfcem. 270:21966; Veldrnan ei al. (1995) 1 Bio'. Chem. 270:16470). Biochemical characteristics of TAIP, such as its molecular wa.jj it and its tendency for dirnerization suggested the possibility that TAB4 may be analogous 1:0 PSGL-1. To investigate the relationship between these two antigens, the foLo iving were tested; 3) whether the antigen precipitated by TAB4 can be recognized by ii: ■jo;runercial'Iy-a.vailable and-PSGL1 antibody; and 2) wheiher an anti-PSGLl anciboc.y can deplete TAB4 from the cell lysate. RL^i T cells ware lysed at a density of 1.0 x 10s cells/ml in lysis buffer (1% Tritr:-iX-1Q0, 20 raM Tris-HCl, pH 8.0,160 roM Nad, 1 mM CaCI*) containing compete protease inhibitor cocktail for i hour, and insoluble material was pelleted at I0.0u0 x g for 10 minutes. These and all subsequent steps were perfomied ai 4°C. The lysate corresponding to 5.0 x 101 cells was incubated with 20 ul of protein G-Sepliarose prc:!cfg.c!ed with 10 ug of anti-PSGL-1 mAb (clone 2PE1, PharMSngeti- San Diego, CA), anti-TAlP mAb, TAB4, or IgG from normal hamster serum. After incubation for 4 hcu:cs at 4'3C, the beads were washed five times with washing buffer (0.05% Triton X-100, 50 HUM Tris-HCl, pH 8.5,400 mM NaCl, 1 JDM CaCl2( 1 xog/ml ovalbumin), and twice v/:ith a similar washing buffer, containing 250 rotoiinstead of 400 n-M NaCl , Bound proteins were eluted with 40 ul of IxSDS sample buffer. Ehted proteins were separated by 6 % SDS-PAGE and transferred to a nitrocellulose ineaabrfriG. The mercibrsnes were hninunoblotted with anti-PSGL-1. mAb, and revealed by peroxidase-conjugated goat anti-rat IgG (H-fL) followed by chenulununcscence (Renaissance, NEN). Surface biotinylated flLtf 1 T cells were lyaed at a density of 1.0 x 10B cells/ml ;tn lyei'fi buffer. The cell extract was incubated with 20 ug of antibody bound to 40 ul of profit G-Sephaiose at 4°C overnight. Depletions were done with anti-PSGL-1 raAb (2PK I) or control ra: IgG, with TAS4 or control normal hamster sensm> Depleted lysa'^s were further .subjected to do inrmunoprecipitation with TAB4 cr anti-PSGL-1 icA.b, respectively. Iniinunotirscipfcates were separated on 6% SDS-polyacrylamide gel ma detected by fluorography. As shown in Fig. 6, anti-PSGL-1 antibody can deplete TAIP protein from T cell lysites. In addition, protein* immunoprecipitated with anti-TA:P antibody (TAB4) can be recognized by anti-PSGL-1 antibody by western amrJysiii. £y -iiDLe .10: Induction of Apfiptosis in Human T Crihby an anti-gStJL-1 Antibody To determine the role, played by PSGL-1 in the apoptosis of human T cells, tin:.ii course experiments we;:e earned out to investigate when activated riutnan T cells acq'jur* sensitivity toward PSGL-I*mediated apoptotic signals, Human T cells' were stimulated with pbytohenjagglutinin (PHA) niirogen and further expanded in IL-2-contakring medium. Activated T cells were harvested and then challenged with anti-PSGL-1 in the presence of IL-2 and cross-linking antibodies. Human peripheral blood was taken from healthy adults, heprini2.ed, and etr'.uhcd for peripheral blood a^ononuclear cells (PEMO) based on differential density usiivg Ficoll-PaqL'.e Plus (Pharmacia Biotech). The PBMC were activate with 1% PHA (Lih Technologies, GibcoBRL) for 48 hour.* and subsequently maintained in 3$wn;binani him:ari IL-2 (5 ng/ml) through the assay period. To assess the apoptosis -inducing ability an anti-human PSGL-1 antibody, the activated cells were treated with; (1) I -tig /ml of the aatMPSGL-1 antibody clone KPL-1 (BD PharMingea) plus cross-licitec rabbit anti-mouse Ig (0.5 ug /ml) (Jackson taj&unoResearchLaboratories); (2) isotype control purified mouse Ig plus cross-linker rabbit aiiti-mouse Ig; or (3) cross-linka: rabbit anti-mouse Ig done. After six hours of treatment, the percentage of early apopmtic cells was determined by FACS, staining with anti-Aimexin V (BD Plmt'Ivliagen) and PI (Sigma). A,5 shown in Fig. 8, signaling triggered by FSGL-1 using an anti-PSGL-1 antbc; dy plus the crosslinks triggered significant level of apoptosis m PHA-activatec frjman PBMC (nifcinly T cells). The percentage of apoptotic cells increased from 8.5'H c:' days 3 to 24% on day 8 in sati-PSGLl treated cultures. Neither isotopio-matched central, nor the cross-linking antibodies alone, had any effect on these cells. fr;c-;irrrpte. 11: Use of antirP;iGL-l Agonist Antibody .o Treat Autoimmune Disease Non-obe=se diabetic (NOD) mice, a well-accepted autoimmune diabetes animal, we::u bred under standard conditions. Spontaneous diabetes developed in the NOD i:.ri as at the age of about 20 weeks. In the experimental group, the mice received three d:)W!; of anti-PSGL-1 antibody (TAB4) intraperitoneally at 300 |xg per mouse at age of ] 1, 13 and 17 weeks. Two additional injections with the same dose were given at the .;..!£ :*;. of 24 and 26 weeks. The control group was given liatnster Ig at the same doss, M'.ci were monitored for glucose aria by Mali-Test Glucose strips (Macherey-Nagel, Q^xt iany) twice every weak after the aae of 15 weeks. Non-fasting urine glucose lovels over 300 rog/d] for two consecutive measurements were considered diabetic. As shown in Fig, 9, TAB4 (anti-PSGL-1) antibody treatment yielded significant jimtccticvri as compared with control antibody treatment. Thus an anti-PSGL-1 i?.r...tibody treatment can dampen the activity of autoimmune T cells and delay the onset ■:i'i;y;?ej!-diabei:es. Other Embodiments It is to be understood that, while the invention has been described in conjunction with the. detailed description thereof, the foregoing description is intended to illustrate aiic not limit Hie scope of tlie invention. Other aspects, advantages, s.nd modifications o;:' tlie invention are within the scope of the claims eer forth below. What is claimed is: r 1. A method of preventing or reducing a T cell-mediated immme response in at. ^-dividual, the method comprising: selecting an individual diagnosed as having or as being at risk of acquiring a ccnLciiti'on characterized by an excessive or unwanted T cell-mediated immune response; and administering to the individual a compound That binds to P-Sekotin G';;y;:opro%eiti Lijjand-1 (PSGL-i) on the surface of a T ceil, wherein the binding of die compound to PSGL-1 on the surface of die T cell indices a signal transduction pathway thff results in the? death of the T cell, thereby preventing or reducbg ,\ T cell-mediated imrmae iesponse in the individual, 2. The method of claim 1, wherein the compound is an antibody or antigen biding fragment thereof that specifically binds to PSGL-1. 3. The raefchod of claim 1, v/herein the compound is a monoclonal antibody that s:?tc:fically binds to PSGL-1. 4. The method of claim 3. further comprising administering an agent that binds ic the? monoclonal antibody and induces the cross-linking of a plurality of PSGL-1 ;m;(j;j:;;;ns on the surface of the T cell. 5. The method of claim 1, wherein the method comprises inducing the cross-Jkiiag of a plurality of PSGL-1 antigens on the surface of the T cell, wherein the <: cuss linking induces the signal transduction pathway that results in death of t cell> 6. The method of claim 1, comprising selecting an individual diagnosed as having an autoimmune disease, 7. The method of claim 1. comprising fcelsoting an individual that has received or k expected la receive an allogeneic or xenogeneic transplant. 8. The method of claim 1, comprising selecting an individual diagnosed ns having an allergic disease. 9. The method of claim 1; comprising selecting an individual diagnosed as hs-v ii{;; a T cell cancer. 10. The method of claim 1, wherein the T ceil is an activated T cell, 11. The method of claim 1, wherein the T cell is a CD4^ T call. 12. The method of claim 1, wherein the T cell is a CDS* T cell. 13. The method of claim 1, wherein the method comprises detecting the in .nib sr of T ceHs in a first biological sample taken from the individual before the ^ministration of the compound and comparing the rtsnlU with the number of T cells in h r;:eond biologies! sample taken from the individual after tlie administration of the VA,: impound. 14. The method of claim 1, wherein the method comprises detecting a biological activity of T cells in a first biological sample taken, from iJae individual l"*?ots the admioistration of the compound and comparing the resulfs with the biological activity of T cells in a second biological sample taken from, the individual ai!er the administration of the compound* 15. The method of claim 1, wherein the administration results in the depletion of at least 20% of peripheral blood CD3+ cells in the individual, 16. The method of cJaim 1 wherein the antibody or antigen binding fragment .:i:;::t:of induces the death of at least 20% of peripheral blood CD3i- cells in the individual aftei exposure to the antibody or antigen binding fragment thereof. L7. A method of :indncing the death of a T cell or a natural killer (NK) cell, the Tied tod comprising; providing a T cell or NK cell expressing FSGL-1 on its cell surface; and contacting flic T cell or MK coll with a compound that binds to PSGL-1 on the sudfceo of the T cell or NK cell, vvhetein the binding of the compound to PSGL-1 on the .-ittrface of the T ceil or NK cell induces a signal traduction pathway that results in tha death of the T cell or NK cell. IS. The method of claim 17, wherein the compound is an antibody or antigen buidfag fragment thereof that specifically binds to FSGL-i. IS, The method of claim 17, wherein the compound is a monoclonal antibody that q:»eciScally binds to PSGL-1, 20. The method of claim 19. further corn-prising contacting the monoclonal antibody with an agent that binds to the monoclonal antibody and induces the cross-linl;k;g of a plurality of PSGL-1 antigens on the suarfitce of the T cell or NK cell. 21. The method of c.ahn 17, wherein the method comprises inducing the cro^s-]:.nkii,g of a plurality of PSGL-1 andgea? on the surface of the T cell, or NK cell, v, [ sriAi iie cross-linking induces the signal transduction pathway thai: results in the <:i of the t csll or nk cell.> 22. The method of claim 17, wherein thfc cell is an activated T cell 23. The method of claim 17, wherein ths cell is a CD4+ T cell. 24. Th« method of claim 17, wherein the cell is a CDS-r T cell. 25. The method of claim 17, wherein, the method comprises assessing the viability of the T cell or 1\K cell after the contacting with the compound 26. The method of cleim 17, wherein the method comprises assessing a ':iojogical activity of the T cell orMC cell after the contacting with the compound. 27. A method of screeoing for a modulator of PSGL-1 function, the method comprising; providing a sell expressing PSGL-1 oh the surface of the cell; contacting the ceil with a test substance; and measuring the viability of the cell after contacting the cell with the- test substance to thereby determine if the test substance is a modulator of'PSGL-1 fhnetioL 28. The method of claim 27, further comprising detecting the daath of the cell incbicc-id by the test substance to thereby determine that Hie test substance is a modulate of I'SGL-I function. 29. The snethod of claim 28, wherein the test substance is an antibody or anUgen binding fragment thereof that specifically binds to PSGL-1. 30. The method of claim 28, wherein the test substance ij a monoclonal actibody that specifically binds to PSGL-1. 31. Thvi method of claim 30> further comprising contacting the monoclonal *Hiu;>::>dy with an agent that binds to the monoclonal antibody \\sxd induces the cross-Ibking of a plurality of PSGL-1 antigens on the sarface of the cell. 32. The method of claim 28, wherein the method comprises inducing the cross-Jirkmg of a plurality of PSGL-1 'Antigens on the surface of the cell, wherein the cross-Imidug induces the signal transduction pathway that results hi the death of the cell 33. The method of claim 28, wherein ths cell is an activate*:. T cell, 34. Th 35. The method cf claim 28, wherein the ceil is a CDS* T cell. 36. The method of claim 23, further comprising manufacturing bulk quantities of 'ii.« test substance and formulating the test substance in apharaiaceutieally acceptable carrier, 37. A kit comprising; a compound that birds to PSGL4 on the surface of a T cell, wherein the bintikig of the compound to PSGL-1 on the surface of the T cell induces a signal transduction pathway that results in the death of the T cell; and instructions for use of the compound to treat autoimmunity, transplant rejection, an ftllargic condition, or aT cell cancer, A method of preventing or reducing a T cell-mediated immune response in an individual substantially as hereinabove described with reference to the accompanying drawings. A kit substantially as hereinabove described with reference to the accompanying drawings.

Documents:

0452-chenp-2004 abstract-duplicate.pdf

0452-chenp-2004 claims-duplicate.pdf

0452-chenp-2004 descripition(completed)-duplicate.pdf

452-chenp-2004-correspondnece-others.pdf

452-chenp-2004-correspondnece-po.pdf

452-chenp-2004-description(complete).pdf

452-chenp-2004-drawings.pdf

452-chenp-2004-form 1.pdf

452-chenp-2004-form 3.pdf

452-chenp-2004-form 5.pdf

452-chenp-2004-form18.pdf

452-chenp-2004-pct.pdf