Chimerism of multiple monoclonal antibodies expressed in a single plant

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 (mAb(M)C) and human anti-rabies mAb 57 (mAb(H)R), 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 (mAb(M)C) and human anti-rabies mAb 57 (mAb(H)R), respectively, were crossed to reproduce F-1 transgenic plant, expressing both mAbs. PCR and immunoblot analyses demonstrated that heavy (HC) and light chain (LC) genes of mAb(M)C and mAb(H)R were present, and that both mAbs were expressed in F-1 transgenic lines, respectively. Quantitative immunoblot for purified mAb also showed the presence of both mAbs in F-1 transgenic lines. However, Cell ELISA analysis showed that in mAb(P)C and mAb(P)R purified from the F-1 transgenic lines (mAb(P)CxR), the binding activity to SW948 human colorectal carcinoma cells was lower than mAb(M)C, and in vitro mAb(M)C, mixed with mAb(H)R (mAb(MH)C+R). The in vitro rabies virus neutralization assay demonstrated that the mAb(P)CxR, from the F1 transgenic plants, had lower bioactivity against rabies virus than mAb(H)57, and mAb(MH)C+R. N-glycan structure analysis revealed that mAb(M)C and mAb(H)R 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 mAb(P)C and mAb(P)R; 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.