Plant-derived EpCAM-Fc fusion proteins induce in vivo immune response to produce IgGs inhibiting invasion and migration of colorectal cancer cells.

Abstract

Key message: Transgenic tobacco plant expressed EpCAM-Fc fusion proteins to induce in vivo immune responses producing anti-EpCAM antibodies inhibiting human colorectal cancer cell invasion and migration. Plant is emerging as a promising alternative to produce valuable immunotherapeutic vaccines. In this study, we examined the in vivo anti-cancer efficacy of epidermal cell adhesion molecule (EpCAM)-Fc and EpCAM-FcK fusion proteins produced in transgenic plants as colorectal cancer vaccine candidates. Mice were injected with plant-derived EpCAM-Fc (EpCAM-Fc^P) and EpCAM-Fc^P tagged with KDEL (ER retention signal) (EpCAM-FcK^P), using mammalian-derived EpCAM-Fc (EpCAM-Fc^M) as positive control. Total IgGs from the immunized mice were used to assess immune responses. ELISA tests revealed that IgGs from mice immunized with EpCAM-FcK^P (EpCAM-FcK^P IgG) exhibited the highest absorbance value for binding affinity to recombinant EpCAM-Fc^M compared to IgGs from mice immunized with EpCAM-Fc^P (EpCAM-Fc^P IgG) and EpCAM-Fc^M (EpCAM-Fc^M IgG). Bio-layer interferometry revealed that EpCAM-FcK^P IgG had a higher affinity value than EpCAM-Fc^M IgG and EpCAM-Fc^P IgG. Cell ELISA revealed that EpCAM-FcK^P IgG exhibited the highest binding activity to EpCAM-positive cells SW480 and SW620 compared to EpCAM-Fc^P IgG, EpCAM-Fc^M IgG, and anti-EpCAM mAb. In the transwell invasion assay, EpCAM-FcK^P IgG significantly decreased the numbers of invaded SW480 and SW620 cells compared to EpCAM-Fc^P IgG, whereas EpCAM-Fc^M IgG had similar numbers. In the wound healing assay, EpCAM-FcK^P IgG showed higher migration inhibition compared to EpCAM-Fc^P IgG in both cell types, with similar results to EpCAM-Fc^M IgG in SW620 cells. These results confirm the applicability of plant systems to produce EpCAM-Fc vaccine candidates, inducing the production of anti-EpCAM IgGs against colorectal cancer cells.