cytoskeleton rearrangement of DCs. In addition to databrought by FACS and ELISA analysis, these findings reinforcethe capacity of TX527, to silence some important features ofDCs that make them potent antigen presenting cells,inducing a more tolerogenic state.

doi:10.1016/j.clim.2008.03.325

Sa.105. Immunomodulatory Effects of TX527, a LessCalcemic Vitamin D Analog, on Human SynchronizedT Cells: a Proteomic ApproachFemke Baeke, Gabriela Ferreira, Evelyne van Etten, LutOverbergh, Chantal Mathieu. Catholic University of Leuven,Leuven, Belgium

The active form of vitamin D3, 1,25-Dihydroxyvitamin D3(1,25(OH)2D3), and certain analogs such as TX527 are potentimmunomodulators. Direct effects of 1,25(OH)2D3 andanalogs on T-cells have been described, but haven't beenfully explored yet. This study aims to investigate theimmunomodulatory effects of TX527 on human T-cells,using a proteomics approach. Since TX527-mediated effectsdepend on vitamin D receptor (VDR)-expression, being onlyexpressed in T-cells upon activation, the benefit of T-cellsynchronization before TX527-treatment was first investi-gated. Human T-cells from healthy volunteers (n=4) wereactivated by anti-CD3+anti-CD28 and treated with TX527(10-8 M) simultaneously (day 0) or on day 2. IFN-γ, IL-10 and24-hydroxylase mRNA expression levels were measured byquantitative real-time RT-PCR at day 4. Strongest effects onIFN-γ, IL-10 and 24-hydroxylase mRNA-levels were seenwhen TX527-treatment was started on day 2 as compared toTX527-treatment on day 0. Therefore, the above T-cellactivation protocol with TX527 (10-8 M)-addition on day 2was used for proteomics analysis. After 24 hours oftreatment, protein samples were separated in 2 pH-ranges(pH 4-7 and pH 6-9) using 2-dimensional Differential GelElectrophoresis (2D-DIGE). Approximately 3400 proteins weredetected in all groups of comparison. Upon TX527-treatment,53 proteins changed significantly (n=4,pb0,05). Differentiallyexpressed proteins are currently identified by MALDI-TOF/TOF. In conclusion, these data demonstrate the necessity of T-cell activation prior to TX527-treatment for optimal effects.Additionally, identification of the differentially expressedproteins will certainly contribute to a better understanding ofthe direct immunomodulatory effects of TX527 on T-cells,opening new possibilities for therapeutic interventions.

doi:10.1016/j.clim.2008.03.326

Sa.106. Antigenic Peptide Administration Not OnlyReduce the Autoantibody Production but alsothe Frequency of CD138+ Cells in New ZealandBlack MiceYi-Chun Cho,1 Ya-Ken Chen,1 Chao-Lin Liu,2 Chia-rui Shen.11Chang Gung University, Kweishan, Taiwan; 2Min Chi Uni-versity of Technology, Taishan, Taiwan

New Zealand Black (NZB) mice spontaneously developautoimmune hemolytic anemia which is resembled to humanAIHA. The major target of the pathogenic erythrocyteautoantibodies is possibly the anion channel protein band 3as CD4+ T cells of NZB mice were also shown to respond toband 3. Previous studies done by our laboratories showedthat band 3 peptide 861-870 contains a dominant auto-reactive helper T-cell epitope with the ability in vivo tomodulate the course of AIHA in NZB mice. In the presentstudy, we demonstrate that not only the production ofautoantibody but also the frequency of CD138+ cells, namelyplasma cells, could be reduced in NZB animals receivingpeptide treatment. In fact, the B cells, bearing B220+, arealso decreased. Moreover, the elevated proportion of CD4+CD25+FoxP3+ cells was noted in the induction of mucosaltolerance by administrating auto-antigenic peptide to theseanemic NZB animals. It appears that NZB mice with highertiters of erythrocyte autoantibodies harbor less CD4+CD25+cells. Most CD4+CD25+ cells are FoxP3+. The question now isbeing addressed is whether these CD4+CD25+ cells are withdirect effects on the production of erythrocyte autoantibo-dies or on the frequency of CD138+ cells. These findings willbenefit further study of T regulatory cell biology interferingwith B cells, particularly in antibody-based autoimmunediseases.

doi:10.1016/j.clim.2008.03.327

Sa.107. Mice Overexpressing the Truncated 1/4CTLA-4 Isoform Have Hyperactive T Cells andAccumulate Memory Cells Without DevelopingSpontaneous Autoimmune DiseaseSue Liu, Zheng Zhang, Mohamed Oukka, Vijay Kuchroo.Brigham & Women's Hospital & Harvard Medical School,Boston, MA

CTLA-4 is a potent inhibitor of T cell activation, whichfunctions mainly via binding costimulatory molecules fromthe B7 family. However, recent reports show that a CTLA-4isoform that lacks B7 binding capacity, ligand independent-CTLA-4, can still inhibit T cell activation. 1/4 CTLA-4 isanother isoform that lacks the ligand binding domain as wellas the transmembrane domain, but so far the function of thisisoform is not known. To investigate the function of 1/4CTLA-4 in vivo, transgenic mice overexpressing this geneticvariant were generated. The frequency of CD44high memoryT cells in 1/4 CTLA-4 Tg mice was almost double that inwildtype littermates, and as the mice aged, the frequencyfurther increased. Unlike in young mice, 1/4 CTLA-4 Tg micemore than a year old displayed a skewing toward CD4+ Tcells, an increase in activation markers, as well as anincrease in Foxp3+ regulatory Tcells compared with wildtypelittermates. T cells from 1/4 CTLA-4 Tg mice proliferatedmore and produced more cytokines than wild type T cells.Despite this extreme state of T cell activation, the miceshowed no overt signs of disease, possibly due to theincreased proportion of regulatory Tcells. This data suggeststhat the function of 1/4 CTLA-4, unlike full length CTLA-4,may not be to inhibit T cell activation.

S115Abstracts

doi:10.1016/j.clim.2008.03.328

Sa.108. CD47 Controls the In Vivo Proliferation andHomeostasis of Peripheral CD4+CD25+ Foxp3+Regulatory T Cells that Express CD103Vu Quang Van, Jinane Darwiche, Marianne Raymond, SylvieLesage, Salim Bouguermouh, Manuel Rubio, Marika Sarfati.Centre de Recherche de l'université de Montreal (CHUM),Montreal, QC, Canada

Peripheral CD103+Foxp3+ regulatory T cells (Tregs) candevelop both from conventional naïve T cells upon cognateAg delivery under tolerogenic conditions as well as fromthymic-derived, expanded/differentiated natural Tregs. Wehere show that CD47 expression, a marker of self onhematopoieitic cells, selectively regulated CD103+ Foxp3+Treg homeostasis at the steady state. We first observed arapid and progressive augmentation with age in theproportion of effector/memory-like (CD44HighCD62LLow)CD103+ Foxp3+ Tregs in CD47-deficient mice (CD47-/-) ascompared to age-matched wild type littermates. Yet,the proportion of quiescent (CD44LowCD62LHigh) CD103-Foxp3+ Tregs remained stable. We next found an increasedproliferation rate (BrdU incorporation) within theFoxp3+CD47-/-Tregs subpopulation that was restricted tothose expressing CD103. Finally, CD47-/-Tregs displayed anormal suppressive function in vitro and in vivo. We proposethat sustained CD47 expression throughout life is critical toavoid an excessive expansion/accumulation of CD103+Tregsthat may overwhelmingly inhibit pathogen or tumor-specificCTL responses.

doi:10.1016/j.clim.2008.03.329

Sa.109. Allelic and Phenotypic Heterogeneity at theAutoimmune Susceptibility Locus IL2RALisa Maier, Christopher Severson, Philip De Jager, DavidHafler. Harvard Medical School, Boston, MA

In a recent genome wide association scan for MS suscept-ibility genes, themost significant Single Nucleotide Polymorph-ism (SNP) outside the MHC associated with MS was located inthe gene encoding the alpha chain of the IL-2 receptorcomplex, IL2RA. The IL2RA gene has previously been associatedto Type 1 Diabetes (T1D) andGraves Disease,making it a sharedautoimmune susceptibility locus. In this study, we testedwhether, similarly to what was shown for the T1D variants, theMS susceptibility allele rs12722489 was associated with levelsof sIL-2RA. First, we quantitated serum levels of sIL-2RA andfound that they are increased in MS subjects compared tohealthy controls (mean sIL-2RA concentration in healthycontrols=2,022 pg/ml [95% CI 1,852 - 2,192]; mean sIL-2RAconcentrations in MS subjects=2,345 pg/ml [95% CI 2,256-2,435]; P=0.0001), an observation previously reported not onlyin MS but also in other autoimmune diseases, infectiousdiseases and malignancies. Using two independent serumcollections consisting of 70 healthy controls and 300 MS

patients, we then determined that the MS susceptibility alleleat rs12722489 is associatedwith increased levels of sIL-2RA in adose-dependent manner in both healthy controls (P=0.00019)and MS patients (P=0.0217). However, in stark contrast to thefinding for T1D susceptibility alleles, we found that the MSsusceptibility allele is associated with higher levels of sIL-2RA.Thus, MS susceptibility is associated with increased sIL-2RAlevels, while T1D susceptibility is associated with decreasedsIL-2RA levels.

doi:10.1016/j.clim.2008.03.330

Sa.110. Fully Human Monoclonal AntibodiesDirected Against Human LIGHT Effectively SuppressAcute Xenogeneic Graft-versus-host DiseaseMediated by Human T CellsMichael Lyman,1 Chris Lynn,1 Rachel Murray,1 OlgaTurovkaya,2 Enrique Rodriguez,1 Amy Coddington,1 IsaoSerizawa,3 Toshiyuki Tanaka,4 Shinichiro Kato,1 RachelSoloff,1 Steve Granger.1 1Kirin Pharma USA, La Jolla, CA;2La Jolla Institute for Allergy and Immunology, La Jolla, CA;3Kirin Pharma, Co., Ltd., Tokyo, Japan; 4Hyogo University ofHealth Sciences, Kobe, Japan

LIGHT is a ligand of the TNF superfamily (TNFSF14) thatpotentiates T cell mediated immune responses and exhibitspro-inflammatory activity. Like other pro-inflammatory TNFsuperfamily counterparts, aberrant LIGHT signaling has beenlinked to chronic inflammatory and autoimmune pathology insuch diseases as rheumatoid arthritis (RA) and inflammatorybowel disease (IBD). Recent reports also indicate that LIGHTmay play a role in atherosclerosis by regulating plasmalipoprotein concentrations by signaling through the lympho-toxin beta receptor (LTβR) and in graft versus host disease(GvHD) through the LIGHT-herpes virus entry mediator(HVEM) co-stimulatory pathway. We have used transchromo-somic KM mice to generate a panel of antagonistic fullyhuman monoclonal Abs (mAbs) that display high affinity forhuman LIGHT and effectively block binding of LIGHT to bothLTβR and HVEM and block LIGHT-mediated chemokinesecretion in vitro. Our anti-human LIGHT mAbs also show invivo efficacy by suppressing disease in an adoptive transfermodel of acute xenogeneic GvHD (XGvHD). Adoptive transferof human peripheral blood mononuclear cells (PBMC) intoirradiated SCID mice depleted of NK cells results in an acutelethal XGvHD mediated by CD4+ and CD8+ T cells. Prophy-lactic treatment with anti-LIGHT mAbs significantly reducesgross pathology and human Tcell numbers in recipient mice.Our data suggest that mAbs against human LIGHT may beefficacious in the treatment of inflammatory diseases of theintestines as well as in the treatment of other T-cellmediated inflammatory diseases.

doi:10.1016/j.clim.2008.03.331

Sa.111. The Human FOXP3+ Regulatory T CellPopulation: Single-cell Study Reveals a FunctionalHeterogeneity

S116 Abstracts