STUDYING DESIGN PRINCIPLES FOR BISPECIFIC ANTIBODIES – THE H-L PAIRING CHALLENGE

Bispecific antibodies (bsAbs) are antibodies with two binding sites directed at different antigens, enabling therapeutic strategies not possible with conventional monoclonal antibodies (mAbs). Since bispecific antibodies are regarded as promising therapeutic agents, several bispecific design modalities have been evaluated, but as most of them are small recombinant fragments, their utility is limited. In a nutshell, the field prefers IgGs.
Two criteria should be met to make bispecific IgGs; one is that each heavy chain will only pair with the heavy chain of the second specificity and that homodimerization be prevented. The second is that each heavy chain will only pair with the light chain of its own specificity and not with the light chain of the second specificity. The first solution to the first criterion (knobs into holes, KIH) was presented in 1996 by a group from Genentech. Additional solutions were presented more recently. However, until recently, no solutions for the second criterion that make it possible to produce a bispecific IgG by an expressing cell were suggested.
We present a solution for the second criterion; an engineered disulfide bond between the antibodies’ variable domains. This approach termed disulfide stabilization, replaces the natural disulfide bond between the CH1 and CL domains. By combining KIH for heavy chains heterodimerization and disulfide stabilization for H-L chain pairing, we efficiently produced several bsAbs in bacteria and in mammalian cells.
In the presentation examples will be provided for the evaluation of some of these bsAbs and future directions of the study will be discussed.