Fully Human Antibody Library, HuPhage
Fully human antibody library constructed by CAT (Cambridge Antibody Technology) or DAYX is prepared as follows: after B cells are isolated from human blood or bone marrow, mRNAs are extracted from the cells. Heavy chain genes composed of VDJ gene segments are synthesized through PCR to make an antibody library. In contrast to the method, PharmAbcine greatly increases the diversity of fully human antibody library by recombination of V gene segments and DJ gene segments when heavy chain genes that are critical for the specificity of antibody are constructed. By that reason, heavy chain gene set of HuPhage can show tens or hundreds of times higher genetic diversity than those of CAT or DAYX.
Preparation of HuPhage Library
The HuPhage Library was completed by inserting a HuPhage heavy chain gene set to the Phagemid for scFv (single chain variable fragment) in which a HuPhage light chain gene set was inserted by using proper restriction enzymes. M13 phage was used as a backbonefor HuPhage Library and the scFvs encoding human-derived antibody fragments were displayed in the ends of M13 Phage to allowitself to bind to specific antigens.
By transforming the HuPhage Library gene set into E-coli, PharmAbcine obtained the HuPhage scFv Display Library with greaterdiversity than 1011 (10E11).
The HuPhage scFv Display Library is a fully human antibody library constructed by assembling naive cDNAs derived from B cells and the antibodies discovered and developed here have the same amino acid sequences as human antibodies. It can reduce the immunogenicity caused by immune responses in the body. As the diversity of scFv in the library is greater than 1011 (10E11), it is assumed that antibodies which can bind to most of the antigens are present in the library. We are discovering and developing the best candidates for antibody therapeutics through our innovative biopanning methods.
The process and technique for screening hit antibody for a specific antigen from the HuPage scFv Display Library is called biopanning. This technique is used to select scFv-displayed phages that are uniquely bonded to antigen. Assuming the fact, the antibody selected through biopanning will become a candidate for antibody treatment that inhibits the function of disease inducer. A simple schematic diagram of biopanning is shown below.
As shown in the diagram, the HuPage scFv Display Library with a diversity greater than 1011 (10E11) is sprayed onto the plate and the plate is washed away with a solvent, the stronger the binding force of the antigen, the more combined with the fixed antigen on the plate. The combined scFv-displayed phage is then eluted using another solvent. The E-coli is transfected to amplify the amount of recovered scFv-displayed phage. Repeating the above procedure three to four times with quantitatively increased scFv-displayed phage through E-coli, gradually increasing the number and intensity of rinsing, the weak binding scFv-displayed phage is washed-out and eventually releases the antigen only the specifically binding scFv-displayed phage can be isolated.
When DNA is isolated from the scFv-displayed phage selected by biopanning, the heavy chain variable region (VH) and light chain variable region (VL) sequences are inserted into the primary antibody backbone to obtain a unique antibody binding only to a specific antibody.
Dual Target Antibody Platform Technology
A dual target antibody is an antibody that can bind to two different antigens at the same time and exert their functions. Considering a conventional antibody therapeutic agent specifically binds to only one antigen and functions, it can simultaneously improve the specificity and binding ability of the target to improve the efficacy by simultaneously binding to two antigens. In addition, if the two targeted antigen have a functional link in relation to the occurrence and outcome of a disease, the synergistic effects of both antigen are expected to be effective by simultaneously hindering the function of the two antigen. It is the next-generation antibody platform technology.
The dual target antibody platform technology of PharmAbcine is based on the candidate material selected through biopanning technique. In other words, the dual targeting antibody is composed of both the selected fully-human antibody candidate, and another one that can exhibit synergistic effects in terms of MOA and connected via a specialized linker to each other. The dual target antibody technology obtained in this way can be classified into DIG-body, PIG-body and TIG-body depending on the manufacturing method.
3G Expression System for High Production Cell Line Development
One production method cannot guarantee the same productivity since production of protein is substantially affected by an inherent structure and properties of the protein. It is important to develop an optimized process for factors affecting protein production. 3G Expression System of PharmAbcine overcomes the limitation in productivity of proteins which are not easily produced.
AVATAR Mouse Model System using Patient-derived Cancer Stem Cells
Professor Nam, Do Hyun, a neurosurgeon at Samsung Medical Center, a founding member of PharmAbcine, created a library of cancerous stem cells derived from 207 brain tumor patients and is using it to evaluate the efficacy of anti tumor therapy. Through this process, we have established a patient-derived tumor animal model system (avatar mouse system) and utilize them to evaluate the efficacy of therapeutic antibody candidaties.
Application of Core Technology