Extended Data Fig. 4: Low affinity anti-CD40 mAb exhibit potent agonism in human systems.
From: Reducing affinity as a strategy to boost immunomodulatory antibody agonism
a, Human DCs were stimulated with various ChiLob 7/4 h2 affinity mutants for 2 days and then evaluated for CD86 expression. The ranking of CD86 MFI was plotted against KD. Means ± SEM, each dot represents an average value from 9 donors. b, Gating strategy and representative histograms for a. c, Human DCs were pre-treated with ChiLob 7/4 h2 affinity mutants for 2 days and co-cultured with allogeneic CD4+ T cells for 5 days. CD4+ T cell proliferation was measured by 3H-thymidine incorporation. Ranking of CD4+ T cell proliferation was plotted against affinity (KD). Means ± SEM, each dot represents the average from 7 donors. d, Human monocyte-derived DCs were pre-treated with ChiLob 7/4 h2 affinity mutants for 2 days and then co-cultured with allogeneic CD4+ T cells at different ratios for 5 days. CD4+ T cell proliferation was measured by 3H-thymidine incorporation. The ranking of CD4+ T cell proliferation was plotted against affinity (KD). Means ± SEM, each dot represents the average value from 7 healthy donors. e, CFSE-labelled PBMCs were stimulated with antigenic peptides and ChiLob 7/4 h2 affinity mutants for 5 days and proliferating CD8+ T cells evaluated for surface expression of CD25. Ranking of CD25 MFI plotted against KD. Means ± SEM, each dot represents the average from 8 donors. f, Gating strategy and representative histograms for CD25 expression of proliferating CD8+ T cells in PBMCs stimulated with CEFX Ultra SuperStim Pool and ChiLob 7/4 h2 affinity mutants. Data representative of 8 donors.
