S42 Abstracts

compared to WT B. fragilis. Ex vivo, CD4+T cells obtainedfrom CLN of mice reconstituted with WT B. fragilis hadsignificantly enhanced rates of conversion into FoxP3+Tregcells and offered greater protection against disease whentransferred in low numbers compared to CD4+T cells fromΔPSA B. fragilis colonized mice. Our results suggest a roleof tolerance induced by commensal components in thecontrol of EAE/MS.

doi:10.1016/j.clim.2009.03.117

OR.103. Induced PLP Peptide-specific RegulatoryT Cells Appear to Suppress ExperimentalAutoimmune Encephalomyelitis in anAntigen-specific MannerHong Zhang, Joseph Podojil, Stephen Miller. NorthwesternUniversity, Chicago, IL

The low number of regulatory T cells (Tregs) and theuncertainty of the requirement of antigen-specificity for theirsuppression present the major clinical challenges of Treg ascellular therapy. Antigen specific Tregs have been reported tobe more efficient than polyclonal Tregs in several diseasemodels. In the present study, Tregs were induced from PLP139-151 specific CD4+CD25-T cells isolated from 5B6 TCR trans-genic mice. To determine if the in vivo suppression of theinduced Tregs (iTregs) was antigen specific, we transferred5B6 iTregs to recipient SJL mice and primed the mice witheither PLP139-151, PLP178-191 or with a combination of thetwo peptides in CFA. The data show that PLP139-151-specificiTregs are able to suppress PLP139-151-induced disease butnot PLP178-191-induced disease. Likewise, transfer of 5B6iTregs to recipient SJL mice is able to inhibit delayed-typehypersensitivity reactions to PLP139-151, but not PLP178-191in mice that were primed with the respective peptides.Importantly, when the 5B6 iTregs were transferred to miceprimed with a mixture of PLP139-151 and PLP178-191, thesemice exhibit a suppressed DTH response to PLP139-151, butnot PLP178-191. In the in vivo proliferation experimentsdetermined by CFSE dilution, CFSE-labeled 5B6-iTregs prolif-erated when transferred to SJL mice followed by priming withPLP139-151, but not PLP178-191. These data indicate that 5B6(PLP139-151-specific) iTregs are re-stimulated when exposedto cognate antigen PLP139-151 under in vivo inflammatoryconditions, however they suppress CD4+effector T cells in anantigen-specific manner. The mechanisms of antigen-specificsuppression are under investigation. This work was supportedby NIH R01 Grant NS026543 and NS048411.

doi:10.1016/j.clim.2009.03.118

OR.104. Induction of Regulatory T Cells by MucosalAdministration of Aryl Hydrocarbon ReceptorLigands and Anti-CD3 Monoclonal Antibody inAnimals and HumansHoward Weiner1, Henry Wu1, Francisco Quintana1, RuthMaron1, Yaron Ilan2. 1Brigham and Women's Hospital,Harvard Medical School, Boston, MA; 2Hadassah Hospital,Jerusalem, Israel

We investigated induction of Tregs in models of auto-immunity and humans by mucosal anti-CD3 antibody andaryl hydrocarbon receptor ligands. Mucosal administrationof proteins is a classic method to induce Tregs. We orally ornasally gave anti-CD3 or Fab2 fragments of anti-CD3 in dosesfrom 0.5ug to 500ug. The optimal dose was 5ug/feeding and0.5ug/nasal. Oral anti-CD3 suppressed EAE given prior to orat disease peak and increased CD4+TGF-b latency associatedpeptide (CD4+LAP+) Tregs in the MLN. Foxp3 was was notincreased by oral anti-CD3. CD4+CD25-LAP+Treg cells sup-pressed in vitro and in vivo in a TGF-b dependent fashion.No modulation of CD3 on the surface of CD4+T cellsoccurred. Oral anti-CD3 suppressed streptozocin induceddiabetes with increased LAP+cells in pancreatic LN. In lupusoral/nasal anti-CD3 suppressed established lupus via an IL-10 dependent Treg (nasal) or TGF-b dependent (oral) Tregcell. In the Ob/Ob mouse model of type 2 diabetes therewas a decrease in the metabolic syndrome and islet/hepaticpathology with induction of TGF-b dependent Tregs anddecreased inflammation in adipose tissue. In humans weadministered oral anti-CD3 to healthy subjects without sideeffects or generation of HAMA responses. In humans, wefound decreased IL-17 production and induction of IL-10 andTGF-b. The optimal dose was 1.0 mg/feeding. Finally, wefound aryl hydrocarbon receptor ligands (which induceFoxp3 Tregs)are biologically active when given orally andsuppress development of EAE by the induction of Foxp3Tregs. These results identify novel and physiologic mechan-isms to induce regulatory T cells via mucosal surfaces that isclinically applicable to a variety of immune mediateddisorders.

doi:10.1016/j.clim.2009.03.119

OR.105. Development of EngineeredAntigen-specific Human Regulatory T Cells for theTreatment of Type 1 Diabetes by TCR Gene TransferTodd Brusko1, Shirley Zhu1, Amy Putnam4, Michael Lee1,Stephanie McClymont1, Weihong Liu1, Michael Nishimura2,Richard Koya3, Timothy Tree4, James Riley5, JeffreyBluestone1. 1University of California, San Francisco, SanFrancisco, CA; 2Medical University of South Carolina,Charleston, SC; 3UCLA School of Medicine, Los Angeles, CA;4King's College School of Medicine at Guy's, King's and StThomas' Hospitals Guy's Hospital, London, United Kingdom;5Abramson Family Cancer Research Institute,University of Pennsylvania, Philadelphia, PA

Current therapies aimed at augmenting Tregs in vivo maybe associated with significant off-target effects. Cellulartherapies using Tregs represent an alternative to systemictreatments, with animal models indicating increasedpotency of antigen-specific Tregs compared to polyclonalTreg populations. However, current technologies are limitedin the capacity to isolate and expand a sufficient quantity ofantigen-specific Tregs for therapy in humans. To overcomethis limitation, we have developed a novel means forgenerating large numbers of antigen-specific Tregs bylentiviral T cell receptor (TCR) gene transfer into in vitroexpanded polyclonal CD4+CD127-/loCD25+T cells. TCR-