Abstract 1626 Cytotoxic T lymphocyte-mediated killing of factor VIII-specific B lymphocytes using a bispecific immunoconjugate

Hemophilia A (HA) is an X-linked recessive bleeding disorder in which patients are deficient in pro-coagulant factor VIII (FVIII) protein. Treatment of HA with regular infusions of FVIII is extremely effective, but 20-30% of patients will develop neutralizing antibodies to FVIII, called inhibitors. The development of inhibitors renders standard treatment ineffective and is a major impediment to therapy. Inhibitors are known to prefer epitopes on the C2 and A2 domains of FVIII, both critical for activity. One approach to tolerize patients to FVIII is to eliminate the population of B lymphocytes that recognize immunogenic epitopes on FVIII. Previous research has utilized cytotoxic T lymphocytes (CTLs) engineered to target FVIII-specific B lymphocytes with a chimeric antigen receptor displaying immunogenic domains of FVIII. As an alternative but related strategy, we have designed and produced a bispecific immunoconjugate to crosslink CTLs with FVIII-specific B lymphocytes. The immunoconjugate engages CTLs with an anti-CD8 arm, and displays the C2 domain of FVIII to attract FVIII-specific B lymphocytes. We hypothesize that contact between the two cell types will initiate CTL-mediated killing. Using C2-specific hybridomas as targets and blood-derived mixed human lymphocytes as a source of T cells, we have demonstrated via B-cell ELISpot assays that the construct is capable of initiating killing. We are continuing to explore the use of the immunoconjugate in tissue culture and are planning to conduct mouse experiments to examine its in vivo efficacy. Supported by NIH grant R21 HL152318 to DWS and JEW.