Extended Data Fig. 6: Development of anti-LILRB3 blocking antibodies for suppressing AML development.
a, Upper: Flow chart of strategy for development of fully humanized antibodies against LILRB3. Lower: The identified antibodies were tested in the LILRB3 chimeric receptor reporter cell assay. b, ELISA results for LILRB3 binders. c, EC50 values of the anti-LILRB3 antibodies based on ELISA. d, Affinities of antibodies #32, #33, #67, and #45 to LILRB3 as determined by Octet. e, Cross-reactivity of the anti-LILRB3 antibodies with LILRAs evaluated with LILRA binding analyses. f, Cross-reactivity of the anti-LILRB3 antibodies with other LILRBs evaluated with LILRB binding analyses. g, Interaction with TRAF2 contributes to the effect of LILRB3 on AML development. CFU assay of MLL-AF9 AML cells expressing wild-type LILRB3 or mutant LILRB3 with mutations disrupting TRAF2 binding (AAA, QEE509-511AAA) or disrupting SHP-1/2 interactions (Y596/626 F) in the presence of control or anti-LILRB3 antibodies (n = 3 independent cell cultures). h, Evaluation of the effect of Fc-mediated effector functions of anti-LILRB3. Upper: Schematic of treatment. Lower: Percentages of GFP + MLL-AF9 mouse AML cells in PB, BM, SPL and LV of mice transplanted with AML cells expressing B3-FL or B3delICD and injected with IgG or anti-LILRB3 #6 (n = 4 independent mice). i, Survival of mice as treated in panel h. j, NF-κB signaling target gene expression (measured by qPCR) in THP-1 cells from BM of xenografted NSG mice treated with anti-LILRB3 #1NA or IgG (n = 3 independent experiments). The data are presented as mean ± s.e.m, and p values were calculated by two-tailed t-test except for i by log-rank test.
