LakePharma HuMAT™ platform leverages its current antibody humanization and affinity maturation capabilities to offer a one-step Antibody Humanization & Affinity Maturation unparalleled in time and costs while delivering consistent high-quality antibody leads.
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1. Antibody sequence analysis and homology modeling of mAb 3D structure.
2. Identification of key positions supporting CDR loop structure and VH-VL interface.
3. Design humanized variants (3VH, 3VL).
4. Assess the humanness of humanized variants by T20 humanness score developed at LakePharma. [Sean H Gao, Kexin Huang, Hua Tu, and Adam S Adler. (2013) Monoclonal antibody humanness score and its applications. BMC Biotechnology, 13:55]
5. Construct and express nine humanized antibodies (each combination of three designed heavy chains and three designed light chains) as well as a chimeric version (rodent variable regions, human constant regions) at the 30 mL scale in HEK293 cells (Tuna293™ Process).
6. Assess expression levels and measure antigen binding by direct ELISA.
7. Measure affinity by SPR, Octet, or competitive binding and compare to chimeric (24401 only).
8. Gene synthesis and molecular construction of four phage chimeras, in Fab or scFv format
9. Fusion of phage chimeras with a large size human light chain library derived from B cells of human donors and screen for higher binding affinity
10. Library screening, affinity measurements, clone selections and antibody purifications, as described under Affinity Maturation
1.Parental antibody VL and VH sequences
2.DNA template (if DNA synthesis is not desired)
1. Top three antibodies based on antigen binding affinity
2. 100 ug purified antibody for the three top clones
3. Study report