Bispecific antibodies (bsAbs) are emerging as the next wave of antibody-based therapies. They combine the specificities of two antibodies and simultaneously bind to two different antigens or epitopes. Advances in genetic engineering technology has resulted in a range of recombinant bispecific antibody formats. However, heterodimeric IgG-like bispecific antibodies, which are based on the heterodimerization of two different IgG molecules, are a promising format because they maintain the overall size and natural structure of human IgG with good stability, half-life, and pharmacokinetic profiles.

There are two chain mis-pairing challenges in the production of heterodimeric IgG-like bispecific maintain. One is to facilitate heterodimerization of two distinct heavy chains and prevent homodimerization of two identical heavy chains. The second is to have correct pairings of cognate heavy and light chains. An efficient strategy to overcome these challenges is to use knobs-into-holes (KIH) technology to prioritize the heterodimerization of two different heavy chains and combine them with a common light chain, based on the generally accepted fact that the affinity and specificity of an antibody are predominantly defined by the heavy chain and can be maintained when an antibody has a non-cognate light chain.

Alphelix applies a versatile approach to directly generate common light chain bispecific antibodies by using yeast surface display of heterodimeric IgG-like bispecific antibodies. In this approach, two heavy chains from existing antibodies to two different antigens are heterodimerized by KIH technology, and introduced into human naïve or synthesized light chain library. Then the resulting bispecific antibody library is displayed on yeast cell surface in heterodimeric IgG-like bispecific antibody format. The library can be simultaneously selected by fluorescence-activated cell sorting (FACS) against two target antigens to isolate common light chains that, in combination with two heavy chains, keep their binding affinity for two antigens. This straightforward approach can efficiently isolate common light chains for various combinations of two existing heavy chains. It generates functional and developable IgG-like bispecific antibodies, which exhibit high affinities in the nanomolar range and have similar biochemical and biophysical properties to parental antibodies.