In the last blog, we learned that the main feature of ADC candidate target antigen is that it needs to be highly expressed on the surface of tumor cells and not expressed or expressed in normal tissues. Next, we will take a look at the characteristics required for the warhead-antibody-of the ADC biological missile.
High Target
The role of antibodies in ADCs is to target specific tumor cells. Therefore, high specificity is the basic requirement to achieve the efficacy of ADC drugs, which can concentrate cytotoxic drugs on the tumor site and avoid toxic effects on normal healthy cells. In addition, the affinity of the antibody to the target antigen will also affect the exertion of the ADC effect. The low affinity is not conducive to the binding of the antibody to the target antigen. However, if the affinity is too high, the antibody is more likely to be removed by the body. Therefore, the affinity of the antibody needs to be appropriate.
Immunogenicity
The immunogenicity of antibodies determines the half-life of ADC in vivo. Early ADC used mouse monoclonal antibodies could induce a strong, acute immune response to in humans (HAMA). The immunogenicity of antibodies determines the half-life of ADC drugs in vivo. Early ADC used mouse monoclonal antibodies to induce a strong, acute immune response to (HAMA) in humans. With the rapid development of cell engineering, antibody engineering and genetic engineering, humanized monoclonal antibodies and all-human monoclonal antibodies gradually replace mouse antibodies, which greatly reduce the immunogenicity of antibodies and effectively prolong the half-life of ADC in blood.
Conjugate Site
High targeting and minimum immunogenicity are the first important factors to be considered in the selection of ADC antibodies. When these two conditions are ideal, try to choose antibodies that contain multiple natural sites that can be conjugated. In clinical trials application of ADCs, IgG molecules is chose, as its not only contain multiple sites, but also facilitate the addition of conjugation sites by modification. In order to maintain the original characteristics of naked antibodies, including half-life and affinity, especially the fixed number of cytotoxins and the edthomogeneity of conjugates, the position and properties of conjugate amino acid residues are very important. The most common conjugate way is through lysine or cysteine. Other functional groups that can be used for conjugate include glutamate, glycosyl, and SNAP-tag. One downside in using the endogenous amino acids for ADC development is a heavily dispersed drug-to-antibody ratio (DAR) caused by the non-uniform distribution of the conjugation sites, especially in the case of Lysine-based conjugations. To achieve a narrower DAR, Creative Biolabs has explored and adopted a variety of methodologies in antibody design and engineering to introduce a series of specific and chemically versatile conjugation sites into the antibody sequences.
Conjugate Strategies
We incorporate Cysteine residues into defined sites on an antibody to create thiol-engineered antibody (EnCys-mAb) and conjugate payload drugs onto those Cysteine residues via thiol-based chemistry.
Unnatural amino acids (UAAs), such as p-Acetyl Phenylalanine, are integrated into antibody sequences via amber stop codon-mediated transcription and their chemically diverse side chains are excellent sites to introduce the payload drug for ADC preparation.
Some enzymes, such as Sortase A, transglutaminases, formylglycin generating enzyme, recognize and modify certain amino acids on a peptide motif to yield chemically activated side chains. Creative Biolabs exploits these approaches and developed Enzyme-mediated modification and conjugation system to help generate ADCs with specific conjugation sites.
Through antibody design, we create a “cave” on the desired locations on an antibody to facilitate meditope trapping and subsequently, achieve site-specific meditope-based conjugations.
We take advantage of the self-splicing inteins and engineer intein-fusion antibodies, thereby achieving C-terminal specific antibody modification and yielding conjugates with DARs at 1 or 2.
The modification of antibody binding sites can directly solve the core problems in the production of ADC, which is conducive to the smooth marketing of drugs after successful development.