Abstract
A variable domain of heavy chain antibody (VHH) has different binding properties than conventional antibodies. Conventional antibodies prefer binding to the convex portion of the antigen, whereas VHHs prefer epitopes, such as crevices and clefts on the antigen. Therefore, developing candidates with the binding characteristics of camelid VHHs is important. Thus, To this end, a synthetic VHH library that reproduces the structural properties of camelid VHHs was constructed. First, the characteristics of VHHs were classified according to the paratope formation based on crystal structure analyses of the complex structures of VHHs and antigens. Then, we classified 330 complementarity-determining region 3 (CDR3) structures of VHHs from the Protein Data Bank (PDB) into three loop structures: Upright, Half-Roll, and Roll. Moreover, these structures depended on the number of amino acid residues within CDR3. Furthermore, in the Upright loops, several amino acid residues in the FR2 are involved in the paratope formation, along with CDR3, suggesting that the FR2 design in the synthetic library is important. A humanized synthetic VHH library, comprising two sub-libraries, Upright and Roll, was constructed and named PharmaLogical. A validation study confirmed that our PharmaLogical library reproduces VHHs with the characteristics of the paratope formation of the camelid VHHs, and shows good performance in VHH screening.
Highlights
In 1993, it was reported that camelids produced unconventional heavy-chain-only antibodies that bind to antigens solely by the variable domain of their heavy chain
We analyzed the constructed variable domain of heavy chain antibody (VHH) structure dataset and found that the structure of the complementarity-determining region 3 (CDR3) loop can be roughly classified into three groups
Binders for the COVID-19 spike protein were isolated from a human VHH library, indicating that for the COVID-19 spike protein were isolated from a human VHH library, indicating that the human VH could be used in the VHH format [45]
Summary
In 1993, it was reported that camelids produced unconventional heavy-chain-only antibodies that bind to antigens solely by the variable domain of their heavy chain. The variable domain of heavy chain of heavy chain antibody (VHH) was expected to be the generation of therapeutic antibodies [1]. In those days, fierce competition in the development of conventional human or humanized antibodies had only begun, and a few pharmaceutical companies were seriously looking at camelid VHHs. Among them, Ablynx (a subsidiary of Sanofi since 2018) generated several VHH clinical candidates, including caplacizumab. Many were surprised when caplacizumab, the world’s first VHH drug, was approved as a treatment for thrombotic thrombocytopenia by the European Medicines Agency in 2018, and by the Food and Drug Administration (FDA). In March 2021, in Japan, a second VHH drug, ozoralizumab, was submitted to the Pharmaceuticals and Medical Devices Agency for approval against rheumatoid arthritis
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