dysplasia, locally invasive tumors, and metastatic disease.Tumor-bearing TRAMP mice revealed systemic decreases iniNKT numbers. Interestingly, the age-dependent increase iniNKT of normal mice was not seen in frankly tumor-bearingTRAMP mice. iNKTwere detectable in the tumor and draininglymph nodes. Uponα-GalCer stimulation, iNKTof TRAMPmiceexhibited a strong Th2 bias both in vitro and in vivo comparedto WT littermates. Since we showed that the latter iNKTdefects are reversible in vitro, we are investigating reversingthese defects in the TRAMP model in vivo. iNKT adoptivetransfer fromWTmice demonstrated establishment of stablecirculating iNKT. Anti-tumor effects are being monitored. Wewill also examine tumor progression in TRAMP mice adop-tively-transferred with in vitro expanded iNKT, analogous toour cancer clinical trial protocol.

doi:10.1016/j.clim.2008.03.094

OR.88. IL-7/IL-7R alpha-Fc Immune Therapy for NonSmall Cell Lung CancerAsa Andersson,1 Anshu Buttan,1 Raj Batra,2 Li Zhu,1 StevenDubinett,1 Sherven Sharmax.1 1University of California LosAngeles, Los Angeles, CA; 2Veteran's Affairs Greater LosAngeles Healthcare System, Los Angeles, CA

T lymphocyte and antigen presenting cell (APC) activitiesare dysregulated in cancer patients. To rescue Tcell activitiesand improve Tcell-APC interactions we are evaluating a chi-mericmolecule in amurine non small cell lung cancer (NSCLC)model. Interleukin 7 (IL-7) is essential for lymphopoiesis, Tcell homeostasis, T cell maintenance, promotion of T cellcytolytic and innate responses. The biological effects of IL-7 ismediated through the high affinity IL-7 receptor (IL-7R)complex that is composed of the ligand binding IL-7Rα chainand the common shared γ chain. Compared to IL-7, IL-7/IL-7Rα-Fc has a longer half life that improves bioavailability andIL-7 mediated T cell stimulation. The Fc portion of themolecule is designed to promote the interactions with APC Fcreceptors. We find that both in vitro and in vivo; IL-7/IL-7Rα-Fc increased proliferation of CD4 and CD8 T lymphocytes,induced high levels of IL-12, IFNγ, CXCL9 and CXCL10 as wellas augmented T cell markers of activation and memory. Theenhancement in these effectors correlated with the anti-tumor benefit of IL-7/IL-7Rα-Fc in an established murineNSCLC in vivo. Our results demonstrate that the IL-7/IL-7Rα-Fc molecule helps to restore and maintain T cell anti tumoractivities in NSCLC, by augmenting T cell activation andmemory phenotype and impacting proinflammatory cyto-kines. This provides a strong rationale for further evaluationof IL-7/IL-7Rα-Fc for the potential translation to lung cancerpatients.

doi:10.1016/j.clim.2008.03.095

OR.89. Domain 4 of ILY Sensitizes Cancer AntibodyTherapy Through Abrogating Human CD59 FunctionWeiguo Hu, Gongxiong Wu, Jose Halperin, Xuebin Qin.Harvard Medical School, Cambridge, MA

CD59 is a key complement regulatory protein that inhibitsassembly of themembrane attack complex through binding toC8 and C9. Complement-dependent cytotoxicity (CDC) iscritical for antibody-based cancer therapy. However, thetherapeutic efficacy of specific anti-cancer antibodies ishampered by the expression of CD59, which is upregulated inmany cancer cells. For example, increased expression ofCD59 is responsible for the resistance of some lymphomas totreatment with Rituximab, a therapeutic monoclonal anti-body against the B-cell specific CD20 antigen. Recognition ofthe role of CD59 in resistance to cancer immunotherapy hasled to the search of a CD59 inhibitor. Intermedilysin (ILY), acytolytic pore-forming toxin secreted by Streptococcusintermedius, lyses human cells exclusively because thetoxin binds to human CD59 (hCD59) with high specificity.Specificity for hCD59 derives from binding of ILY's domain 4(ILY4) to amino acids 42-58 in hCD59, which also participate inbinding to C8 and C9. We hypothesized that truncated ILY4would abrogate hCD59 function and thereby increase anti-body-mediated CDC in cancer cells. We report here thatrecombinant ILY4 (114 residues) expressed in E. coli.,specifically blocks hCD59 function. Treatment of Rituximabresistant B lymphoma cells with ILY4 increased CDC with anIC50≈ 33 nM in vitro. In a nude mouse xenograft model of thesame Rituximab resistant B lymphoma cells, ILY4 (2.5 ug/gbody weight) significantly decreased tumor size without anyapparent toxicity. We conclude that ILY4 may represent aninnovative adjuvant for cancer immunotherapy in generaland for antibody-resistant cancers in particular.

doi:10.1016/j.clim.2008.03.096

OR.90. Characterization of B LymphocytesHarbored by Germ Cell TumorsSimon Willis,1 Kimberly L. Shampain,1 Scott Rodig,2 ScottMallozzi,1 Stefany Almendinger,1 Jeffrey Bruce,3 KevinO'Connor.1 1Harvard Medical School/Brigham & Women'sHosptial, Boston, MA; 2Harvard Medical School/Brigham &Women's Hosptial, Boston, MA; 3Columbia University Collegeof Physicians and Surgeons, New York, NY

Intracranial germ cell tumors (germinomas) are a hetero-geneous group of malignant brain lesions that almostexclusively affect children and young adults. Testiculargerm cell tumors, termed seminomas, comprise 30–40% ofall testicular tumors. Studies performed over twenty yearsago established that many germ cell tumors commonly harboran immune cell infiltrate that includes substantial numbers ofB cells. Yet, little is known about the role of the immunesystem in this tumor family. Thus, to gain a deeper under-standing of the role the B cells play in this tumor tissue, weexamined the molecular characteristics of the B cell antigenreceptor in tumor-associated B cells. Germinomas, semino-mas and control tissues were sectioned then tissue regionsand individual B and plasma cells were isolated through lasercapture micro-dissection. Immunoglobulin gene transcriptswere used to determine the variable region sequences.Analysis of these sequences revealed clear evidence ofaffinity maturation in that the cardinal features of an antigendriven B cell response were present: significant somatic

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mutation, isotype switching, receptor revision and codoninsertion/deletion. Additionally, these data revealed bothclonal expansion and variation, suggesting that local B cellmaturation occurs within the tumor microenvironment and/or local lymph nodes. In contrast, sequences from controltissues and peripheral blood revealed none of thesecharacteristics. Collectively, these data strongly suggestthat an adaptive and specific immune response, mediated byB cells, is occurring within or near the tumor environment.This response is likely to be driven by tumor-associatedantigens.

doi:10.1016/j.clim.2008.03.097

OR.91. The Critical Role of IKDC in the AntitumorEffects Mediated by the Combination Therapy withImatinib and IL-2Evelyn Ullrich, Grégoire Mignot, Mathieu Bonmort, MagaliTerme, Nathalie Chaput, Laurence Zitvogel. InstitutGustave Roussy, Villejuif Cedex, France

Interferon producing killer dendritic cells (IKDC) wereinitially described as tumor infiltrating cells that mediatedthe anti-tumor effects of combination therapy with imatinibmesylate (IM) and IL-2 in B16F10 melanoma-bearing mice(Taieb J. et al. Nat. Med. 2006). IKDC are characterized bytheir distinct surface marker profile (CD11cintB220+NK1.1+)and purified out of spleen, lymph nodes or bone marrowalong those selection criteria. Our recent work leads to thedefinition of IKDC as unique NK cell subset, distinguishablefrom B220-NK cells by their functional dependency on trans-presentation of IL-15 for their ex vivo proliferation, produc-tion of CCL2, TRAIL-dependent high killing capacity and theinduction of antitumoral immune response. Moreover, theantitumor effects of the combination therapy correlatedwith a CCL2-dependent recruitment of IKDC into tumor beds.We showed that IKDC were able to reduce tumor growth inRag-/- x IL-2Rγ-/- mice. Importantly, IKDC are licensed bytumor cells as they produce IFNγ, and up-regulate matura-tion markers (MHC class II, CD86) in the presence of tumorcells. After tumor encounter and killing, IL-15/IL-15Rαtriggered-IKDC can even induce significant protective immu-nity against tumor rechallenge, even in the presence of highconcentrations of immunosuppressive TGF-β. Finally, ournew data suggest that, in tumoral context, IKDC werecapable to present antigens and to prime T cells in vitro andin vivo. This work will further unravel the detailed mechan-isms of antigen presentation by IKDC. As IKDC seem to be apromising tool for immunotherapy, it is of great interest tocharacterize the human counterpart of IKDC.

doi:10.1016/j.clim.2008.03.098

OR.92. The Damage Associated Molecular PatternMolecule [DAMP] High Mobility Group Box Protein-1[HMGB1] is an Activator of Autophagy DrivingImmunityDaolin Tang, Rui Kang, Adam Farkas, Michael Lotze. Universityof Pittsburgh School of Medicine, Pittsburgh, PA

Introduction: The nuclear protein, HMGB1 is linked todiverse extracellular immune roles including release fromnecrotic cells and secretion by activated macrophagesengulfing apoptotic cells. Autophagy is an important cellularresponse to stress or starvation. At present, the immunolo-gical impact of autophagic cell death is not well character-ized. Here we report that HMGB1 is an important activator forautophagy and mediates subsequent immune responses.Methods: H2O2, rapamycin and starvation were used toinduce autophagy. The autophagosome marker LC3 aggrega-tion as well as HMGB1 translocation and release was assessedin HCT116 cancer cells. Murine embryonic fibroblasts (MEFs)obtained from HMGB1+/+ and HMGB1-/- mice were used toassess the role of HMGB1 protein in the cellular response toautophagic stimuli. Western blotting with specific antibodieswas used to assess signaling. Results: Cell viability experi-ments with H2O2, rapamycin and starvation demonstratedthat HMGB1-/- MEFs were more resistant to these treat-ments when compared with HMGB1+/+ MEFs. HMGB1-deficient cells had decreased LC3 aggregation followingtreatment with autophagic stimuli. Phosphorylation of ERKand p38 MAPK after rapamycin treatment, was increased inHMGB1-deficient cells. HCT116 cells released HMGB1 withautophagic stimuli. Conclusions: These findings suggest thatHMGB1 plays a role in modulating autophagy in cancer cells,regulating the response to stress and starvation. Given thecritical emergent role of autophagy in promoting effectiveantigen presentation in myeloid dendritic cells and the use ofsimilar machinery in promoting phagocytosis, strategies toassess autophagy in cancer cells and recruited inflammatorycells in the tumor microenvironment is an important goal.

doi:10.1016/j.clim.2008.03.099

Th17 Cells in InflammationSunday, June 8

2:45 pm–4:45 pm

OR.93. Quantitative Alterations in T Cell Subsets inSystemic Sclerosis (SSc) Patients. Th17 Subset isIncreased While Tregs are Decreased in SScSergio Escobar-Hernández,1 Maria Vargas-Rojas,2 MarianaDíaz-Zamudio,1 Luis Fajardo-Hermosillo,1 JavierCabiedes-Contreras,1 Tatiana Rodríguez-Reyna.1 1InstitutoNacional de Ciencias Médicas y Nutrición Salvador Zubirán,Mexico, Mexico; 2Instituto Nacional de EnfermedadesRespiratorias b Ismael Cosío VillegasQ, Mexico, Mexico

Introduction: SSc is an autoimmune disease character-ized by fibrosis and vasculopathy. A key feature ofinflammatory lesions is the presence of T cells (CD45RO+).High levels of IL-17 have been reported. This studysuggested that the T-cell activity in SSc takes placeoutside the conventional Th1/Th2 subsets. Aim: Toquantify T helper subsets and Tregs in peripheral bloodfrom SSc patients. Methods: Blood samples from 80consecutive patients (n=6 men) with SSc were obtained.Clinical evaluation and organ involvement were determinedusing the Medsger severity scale. Controls were collected from

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