FIGURE 4.
Evobrutinib inhibits B cells and mast cells in vivo and shows a prolonged PD effect despite its short t1/2. (A) Dose-dependent inhibition of B cell activation. Mice were administrated evobrutinib PO at the indicated doses (filled circles). Whole blood was drawn 1 h later and stimulated with anti-IgD or left unstimulated (empty circles). The differences in MFI of the activation marker CD69 (Δ MFI CD69) between stimulated and unstimulated samples is shown, as well as percent inhibition compared with vehicle and plasma exposure. One-way ANOVA followed by Dunnett posttest indicated statistically significant inhibition at 3 mg/kg. (B) Time-dependent inhibition of B cell activation. Mice were dosed PO with 12 mg/kg evobrutinib, and B cells were stimulated ex vivo at the indicated time points. Plasma exposure is indicated. At 16 and 24 h, evobrutinib was nd. (C and D) Time course of BTK occupancy (C) and plasma exposure (D) in DBA/1 mice at different doses. The detection limit for evobrutinib was 0.1 ng/ml. Values below LLOQ were visualized as 0.1 ng/ml in the graph. (E) Dose-dependent inhibition of mast cell degranulation in mice by evobrutinib. Animals were dosed 1 h before the DNP challenge. Six to seven animals per group were used. Percent inhibition relative to the vehicle group is indicated. (F) Time-dependent inhibition of mast cell degranulation. Animals were dosed with 39.5 mg/kg evobrutinib at indicated time points. Percent inhibition is indicated. Statistical analysis was performed as in (E). *p < 0.05, **p < 0.01, ****p < 0.0001, one-way ANOVA with Dunnett posttest for treated groups compared with vehicle. ####p < 0.0001 compared with sham. LLOQ, lower limit of quantification; nd, no longer detectable; PO, per os (orally).