Chimerism of multiple monoclonal antibodies expressed in a single plant

Abstract

Transgenic plants offer a source for the sustainable, safe, and large-scale production of therapeutic recombinant proteins. In this study, both murine anti-colorectal cancer mAb (mAbMC) and human anti-rabies mAb 57 (mAbHR), expressed in a single plant were investigated for their cancer cell binding activity and rabies virus neutralization activity, respectively. Transgenic plants, expressing murine anti-colorectal cancer mAb CO17-1A (mAbMC) and human anti-rabies mAb 57 (mAbHR), respectively, were crossed to reproduce F1 transgenic plant, expressing both mAbs. PCR and immunoblot analyses demonstrated that heavy (HC) and light chain (LC) genes of mAbMC and mAbHR were present, and that both mAbs were expressed in F1 transgenic lines, respectively. Quantitative immunoblot for purified mAb also showed the presence of both mAbs in F1 transgenic lines. However, Cell ELISA analysis showed that in mAbPC and mAbPR purified from the F1 transgenic lines (mAbPC×R), the binding activity to SW948 human colorectal carcinoma cells was lower than mAbMC, and in vitro mAbMC, mixed with mAbHR (mAbMHC+R). The in vitro rabies virus neutralization assay demonstrated that the mAbPC×R, from the F1 transgenic plants, had lower bioactivity against rabies virus than mAbH57, and mAbMHC+R. N-glycan structure analysis revealed that mAbMC and mAbHR had Golgi type (94 and 14%) and ER type (6 and 86%), respectively, and the purified mAbs from the F1 transgenic plants had Golgi type (75%) and ER type (25%). These results indicate that the F1 transgenic plant produced both mAbPC and mAbPR; however, the HC and LC proteins of each anti-rabies virus and anti-colorectal cancer mAbs were assembled randomly, resulting in chimerism in HC and LC assembly for mAb.