In recent years, therapeutic antibody have developed rapidly, and have achieved remarkable effects in the fields of cancer, autoimmune diseases and organ transplant rejection treatment. Based on the different mechanisms of targets, bispecific antibody that targeting dual targets or epitopes, have once again stimulated people's imagination and enthusiasm for better effect based on multiple functions and forms of antibody drugs.
Because of the complementary and unique mechanisms of bispecific antibody, it is considered to have better therapeutic effect. Then, various bispecific antibody platforms arise for application. According to incomplete statistics, there are currently more than 50 bispecific antibody platforms that display various forms of bispecific antibody molecules. But this also brings a lot of problems, such as the risk of drug-ability of different forms of bispecific antibodies, the difficulty of production process, the expression level of antibodies, the correct light and heavy chain pairing, the correct pairing of different heavy chains and so on.
Common light chain bispecific antibody, as a type of various bispecific antibodies, have very significant advantages. Firstly, the structure of common light chain bispecific antibody are similar to natural IgG format, which will reduce the risk on drug-ability. Secondly, because the antibody heavy chains against the two targets share the same light chain, there is no light chain mismatch with heavy chain. Finally, by using the “Knobs into holes” technology and the same charge-repelling technology, it almost avoids homo-type heavy chain mismatches, allowing the common light chain bispecific antibody to achieve a purity of over 95%. Therefore, common light chain bispecific antibody have great advantages on drug-ability and antibody production process control. Based on the above advantages of the common light chain antibody, Sanyou has constructed a common light chain library platform for bispecific antibody discovery.
Based on representative human light chain germline sequence and human heavy chain sequence derived from tens of billions of PBMCs with extremely high diversity, Sanyou’s common light chain library was constructed with ten billion library size. At present, a semi-synthetic common light chain antibody library, in which the heavy chain CDR3 are synthetic based on heavy chains from tens of billions of PBMCs, is also under construction, it will further expand the diversity of heavy chain library.
Dozens of binders could be acquired for different targets by screening sanyou's common light chain library and the library could meet the needs of customers to screen against different epitopes on the same target or different targets. Moreover, obtained common light chain antibodies are human naïve antibodies, which provides the “raw material molecules ” for bispecific antibody development.
Acquired binders for different targets have the same light chain, which could be constructed to bispecific antibody directly.
The structure of common light chain bispecific antibody are similar to natural human IgG format, which shows advantage on drug-ability and production process.
Acquired common light chain antibody are human naïve sequence.
Dozens of binders for different targets could be acquired from sanyou’s common light chain library.
Light chain design -- Sanyou's common light chain library were designed by referring to the characteristics of humanized mouse light chain germline and germline of therapeutic antibodies.
Naïve heavy chain -- Derived from tens of billions of PBMCs with extremely high diversity.
Semi-synthetic heavy chain – synthetic CDR-H3 based on amino acid composition of heavy chains from tens of billions of PBMCs.
The heavy chain germline distribution is specially designed - heavy chain gene family distribution in common light chain library is consistent with the distribution in naïve human body.
Overview of library construction
Sanyou's common light chain library platform has 3 libraries based on a unique light chain sequence from IGKV1 or IGKV3 gene family, respectively, and its heavy chain sequence derived from tens of billions of PBMCs.
Table on the right displayed the data of library size.
Diversity of common light chain library
Analysis of heavy chain diversity–length distribution of CDR-H3
As shown in Fig. 1 the length distribution of human heavy chain CDR3 from tens of billions of PBMCs. By sequencing and analyzing about 200 heavy chains, it show that all sequences are unique. By analyzing the length distribution of CDR3, it is highly in consistent with that reported in the IMGT database. These data all indicate the extremely high diversity of heavy chain sequences in the Sanyou's common light chain library.
Fig. 1 Length distribution frequency of CDR-H3
Analysis of heavy chain gene family distribution of three common light chain libraries
As shown in Fig. 2, germline distribution of heavy chain in three common light chain libraries show highly consistency with that in naïve human body.
Heavy chain germline distribution in naïve human body
Fig. 3 shows the distribution statistics of the heavy chain germlines in the naïve human body. Data is from reference below.
Tiller, T., et al. (2013). "A fully synthetic human Fab antibody library based on fixed VH/VL framework pairings with favorable biophysical properties." MAbs 5(3): 445-470.
Fig. 3 The distribution statistics of the heavy chain
germlines in the naïve human body
One Case for Common light chain library screening
In this case, HER2 was selected as a representative target to screen for common light chain antibodies against the HER2 double epitope (ECD2 and ECD4). A total of 24 common light chain antibodies were obtained.
Competition assay will be carried out with reference antibody trastuzumab and pertuzumab to ensure at least 2 or more recognition epitopes are acquired.
The following graph is the affinity ranking data of the obtained 24 antibodies.