Tolerance induction by fixed, Achr-targeted, antigen presenting B cells

115 TOLERANCE INDUCTION BY FIXED, ACHR-TARGETED, ANTIGEN PRESENTING B CELLS Johannes Reim, M.D., Kevin R. Mclntosh, Ph.D. Daniel B. Drachman, M.D., Johns Hopkins University, School of Medicine, Baltimore, Maryland 21205, USA The pathogenesis of myasthemia gravis involves a T cell dependent, antibody-mediated autoimmune response directed against acetylcholine receptors (AChRs). Inactivation of AChR-specific T cells should interrupt the immune res~x~e, resulting in therapeutic benefit. These T-ceils have receptors for processed antigen (AChR) in the context of the appropriate MHC Class H. In order to target these T cells, we used specially modified syngeneic B cells to serve as AChR Antigen Presenting Cells (AChR-AP~). To accomplish this, we bound torpedo AChR to surface immunoglob',:!in (slg) of B cells, using heterobifunctional conjugates consisting of antibodies directed against B cell-sIg and monoclonal antibody directed against ACb.R~B cells treated with conjugate plus AChR processed and presented AChR effectively, as evidenced by vigorous lymphoproliferatiw responses of purified T cells from AChR-primed Lewis rats and of AChR-specific T cell lines. "Anergy" of T cells can be produced by incompletely stimulating them with fixed APCs. In this study, T cell unresponsiveness was induced by culturing AChR-specific T cells with pamfurmaldehyde-fixed AChR + conjugate-pulsed B cells. After 20 has of coculture, restimulation of the T cells with fresh APC plus AChR resulted in a markedly reduced proliferative response. The theoretical and practical therapeutic implications of these results will be discussed.

BINDING OF ACETYLCHOLINE RECEPTOR ct-SUBUN!T PEVrIDES "re HLA-A2

F. BaggilJ, J. Eivin 3, A. Townsend 3, R. Mantegazza 2, A. Vincent 1, j . Newsom.Davis 1. l Neurosciences Group and 3 Molecular Immunology Group, Inst. Molecular Medicine', Oxford (UK) 2 Dept. Neuromuscular Disease, "C.Besta" Neurological Institute, Milan (I). Acetyicholine Receptor (AChR) specific autoantibodies and class-lI restricted CD4+ T lymphocytes can be detected in myasthema gravis (biG) patients, but the HLA haplotypes A1-B8-DR3 and A3-B7-DR2 are found with increased frequency in MG, suggesting that class-I molecules could also be involved in disease susceptibility. Class-I specific peptides can support the assembly of class-I molecules in vitro, stabilizing a conformational change in heavy chain and its association with p2-microglobulin (1). Thus, binding of peptides can be detected by immunopreeipitation of HLA-A2 molecules from the mutant human cell line,. 174/T2, using a confoxmational dependent mAb (2). 44 synthetic pep,ides, corresponding to the human AChR a-subunit, including also those distinguishing between the two a-subunit isoforms (-P3A and +P3A)(3), have been studied. Several A2-binding peptides were identified in the extracellular portion (p87-103, p108-129, p157-188), in the cytoplasmic region (p310-327, p391-417) and across the junction between the products of exon P3A (p') end P4 (p' 17'-25'/59-65). (1) A. Townsend et ai., Cell 62:285-295, 1990. (2) V. Cermldolo et al., Nature 345:449-452,1990. (3) D. Beeson et al., EMBO J. 9:2101-2106, 1990.