Fig. 1

DB-1310 binds HER3 via a novel epitope with high affinity and internalization capability. (a-c) The binding of DB-1310 and its parental antibody to human HER3, cynomolgus HER3, human HER2, human HER4 and human EGFR was measured by ELISA. (d) MDA-MB-453 cells were stained with 10 nM biotin-labeled patritumab in the presence of DB-1310 or naked patritumab at serial dilutions of up to 500 nM. The binding of biotin-patritumab to cell surface HER3 was detected by streptavidin-PE staining. The Mean fluorescence intensity (MFI) of PE was measured for each condition, and the inhibition rate was calculated as (MFI _{biotin−patritumab only} – MFI _{test condition})/ MFI _{biotin−patritumab only} x 100%. (e) Internalization of pHrodo-labeled DB-1310 or HER3-DXd was measured by an Operetta CLS high-throughput microplate imager. (f) MDA-MB-453 cells were cultured with pHrodo-labeled DB-1310 for 12 h and images were obtained by confocal microscopy. Lysosomes and nuclei were labeled with LysoTracker (green) and Hoechst (blue), respectively