Although significant advances were achieved in clinical immunosuppression in the early 1980's, recently developed immunosuppressive agents such as FK 506 and mycophenolate mofetil have had relatively little impact on the problem of graft rejection. In addition, although several new immunosuppressive monoclonal antibodies (mAbs) have been tested clinically, none has been shown to possess efficacy superior to OKT3. Our laboratory has developed several new mAbs that possess potent T cell inhibition properties. The proposed research focuses on development of the most promising of these mAbs. This mAb, 5H7, is an anti-human Class MHC mAb that differs from other anti-MHC mAbs in that it binds exclusively to a monomorphic determinant of the alpha3 domain. Experience with this mAb has provided the initial evidence that anti-Class I MHC mAbs act directly at the level of the responding T cell in suppressing cellular responses. This mAb is also a potent inducer of apoptosis in peripheral blood lymphocytes, thus confirming previous observations in mice with alpha3 domain-specific anti-Class l mAbs. Recent studies have demonstrated that TCR signaling augments apoptosis mediated by 5H7 mAb. The proposed studies will provide a fundamental understanding of the mechanisms by which this mAb suppresses T cell responses and induces apoptosis in lymphocytes. The effects of 5H7 on T cell signaling with particular attention to early protein phosphorylation events will be determined. The role of accessory cells in 5H7-mediated suppression of T cell responses will be examined by dissecting individual effects of 5H7 binding to T cell class I molecules, accessory cell class I molecules, and accessory cell Fc receptors using digest fragment preparations of 5H7 and control mAb, and specific types of accessory cells. The apoptosis-inducing properties of 5H7 will also be investigated. The sensitivity of individual T cell subsets to 5H7-induced apoptosis will be determined. Other proteins known to regulate apoptosis in lymphocytes (Fas, Fas ligand, Bcl-2, Bcl-x) will be examined for possible participation in 5H7-induced apoptosis. The nature of TCR- mediated enhancement of 5H7-induced apoptosis will be examined by dissecting the role of TCR and class l signaling strength in the induction of apoptosis. T cell subsets will be targeted for in vivo depletion (by combined treatment with 5H7 and anti-Vbeta mAb), including antigen- specific alloreactive T cells (by combined treatment with 5H7 and alloantigen).
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