Figure 2
In vitro biological function characterizations of HX009. (A) Reporter assays for both target engagement. Left panel: Different concentration of HX009 and HX008 activated luciferase reporter expression in reporter cell lines by down-regulating PD1 /PD-L1 interaction, and EC50 was determined according to four-parameter equation fitting curves36; Right panel: CD47-hFc induced activation induced cell-death (AICD) in Jurkat-CSR reporter cells, where reporter cells were incubated with different concentrations of HX009, Magrolimab and hSIRPα-Fc to block CD47-SIRPα interaction to prevent reporter cells from AICD; IC50 was calculated per the four-parameter equation fitting curves. Data were shown as average signal, n = 2. (B) T cell from two different donors were SEB stimulated, followed by reactivation by HX008 and HX009 after. IL-2 (upper panel) and IFN-γ (bottom panel) secretion were measured via an ELISA method. SEB only treatment was also measured for basal cytokine level. Data were shown as average signal, n = 2. p-values: ***:0–0.001, **: 0.001–0.01, *: 0.01–0.05 compared to the SEB-only treatment. (C) Activation of lymphocytes by HX009 treatment in mixed lymphocyte reaction (MLR) assay. IL-2 (left panel) and IFN-γ (right panel) secretion were measured via an ELISA method. Blanks and IgG4 isotype control treatment were also measured for basal cytokine level. Data were shown as average signal, n = 2.
