Targeted delivery of anti-CTLA-4 antibody downregulates T cell function in vitro and in vivo

Abstract CTLA-4 is a T cell surface molecule that binds to the costimulatory molecules CD80 and CD86 on antigen-presenting cells and downregulates T cell function. Therefore, we wanted to test whether antigen-specific activated T cells could be inhibited through directed CTLA-4 signaling using a bispecific antibody (BiAb) capable of simultaneously binding to CTLA-4 and a tissue-specific antigen. The BiAb was prepared by linking two separate monoclonal antibodies against CTLA-4 and the thyroid-stimulating hormone receptor (TSHR). The mouse B cell lymphoma line M12 (H2 d ) was used to induce alloreactive T cells in CBA/J mice (H2 k ); M12 cells stably transfected with the cDNA encoding murine TSHR (mM12) were used to restimulate the alloresponse in vitro. Results of assays for in vitro T cell proliferation, IL-2 production, and cytotoxicity in the presence of BiAb demonstrated that the BiAb could inhibit the T cell alloresponse when stimulated with mM12 cells but not with M12 cells. This effect was dependent on binding of TSHR-bound BiAb to CTLA-4, since the addition of soluble CTLA-4-Ig blocked the inhibitory effect. Injection of mM12 cells, along with the BiAb, not with antibodies against TSHR or CTLA-4 either separately or together, into CBA/J mice (H2 k ) downregulated alloreactive T cell responses. Our study demonstrated that the presence of CTLA-4 signaling molecules on the surface of target cells can protect those cells from immune attack by antigen-specific T cells and suggested that a similar approach could have potential therapeutic value in transplant rejection and tissue-specific autoimmune diseases.