Generation Of Bispecific Antibody Based On "BAPTS" Platform And Pharmacodynamics Study

Bispecific antibody(BsAb)is a kind of recombinant proteins produced by antibody engineering technology.Bispecific antibody could recognize and bind two different epitopes,and play a new biological function.At present,there are three bispecific antibodies approved on the market and over 80 bispecific antibodies in the clinical development.However,there is a series of challenges remained in antibody generation and pharmacodynamics.In this study,CD3×EpCAM and CD3×CD19 bispecific antibodies were successfully generated by our own technology platform“BAPTS”(Bispecific Antibody by Protein Trans-splicing).For“BAPTS”platform,the bispecific antibody was divided into two parts in hinge region and generated by the split intein.One part was conjugated with Fc domain and the other part without Fc domain.Two parts were expressed in two mammalian cells system respectively.Two antibody fragments were conjugated in vitro by protein trans-splicing reaction mediated by the split intein.The“Knobs-into-Holes”or other Fc engineering methods also were adopted in molecular construction to contribute correct pairing of the heavy chains.This technology platform avoided the problems of light chain mismatch and the correct assembly of heavy chain,which could produce IgG1 antibodies close to the natural structure of antibodies.The preliminary pharmacodynamics study was conducted on CD3×EpCAM bispecific antibody.The EpCAM affinity was characterized using biolayer interferometry.Bioactivities were measured by T-cell activation,T cell recruitment and T-cell based cytotoxicity in vitro,and in vivo xenograft tumor inhibition experiments in vivo.The results showed the affinity to EpCAM were 4.46×10-8 M,the IC₅₀ of cytotoxicity to SK-OV-3,SW480 and OVCAR-3 Cell lines were 0.248 ng/m L,2.315 ng/m L and1.129 ng/m L respectively.In vivo bioactivity study,CD3×EpCAM Bs Ab could inhibit xenograft tumor growth.The results indicated that CD3×EpCAM bispecific antibodies could be developed as clinical drug candidates.