Method of generating a synthetic antibody library, said library and application(s) thereof
Inventors
Chatterjee, Sohang • IYER RODRIGUES, Kavitha • Ghosh, Maloy • Maity, Sunit • Unnikrishnan, Divya • MANJUNATH BANGALORE MUNIRAJU, Yogendra • Murugesan, Sathyabalan • Mukunda, Pavithra • Prasad, Bhargav • Kamanagowda, Veeresha • Bhattacharjee, Sanghamitra • KUMAR DAKSHINAMURTHY, Pravin • Halan, Vivek • Srinivasan, Sankaranarayanan • Hora, Anuradha • Natarajan, Bairavabalakumar • Nair, Karthika • Thanigaivel, Aswini • Amol, MALIWALAVE • RAVINDRA SHENOY, Bharath • Bhima Rao, Sahana • PRAKASH CHAKRABARTY, Subhra • KUMAR DUBEY, Ashvini • Khan, Amir • Sharma, Ankurina • Sharma, Rashmi • Tiwari, Anurag • Kumar, Santosh • PATEL, Shivani • MARKANDA, Nikitha
Assignees
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Publication Number
US-11230706-B2
Publication Date
2022-01-25
Expiration Date
Abstract
The present disclosure relates to a method of generating an antibody library, not limiting to a synthetic antibody gene expression library built on pool of consensus nucleic acid sequences by using codon replacement technology. The present disclosure also relates to a synthetic antibody library generated by employing the method of the present disclosure and application(s) of said antibody library.
Core Innovation
The invention provides a synthetic gene expression library encoding first fusion proteins in which each fusion protein comprises a fusion of a display peptide, a human antibody, and an expression tag. The human antibody includes a human antibody variable heavy chain selected from specified SEQ ID NOs, and the variable heavy chain has a CDRH3 region that is modified from a respective SEQ ID NO.
The library encodes modified CDRH3s having lengths from about 4 to about 23 amino acid residues, with amino-acid composition specified by position-dependent X1–Xn sequences and SEQ ID NOs. The invention introduces controlled CDRH3 diversity by defining positional amino-acid frequency and positional correlation, and by using Shannon-entropy based true diversity, including measures of richness and abundance.
The synthetic library is generated from consensus heavy/light chain scaffolds and uses codon replacement to introduce the modified CDRH3 diversity within the human antibody variable heavy chain. The invention also supports antibody formats that include Fab and ScFv associated with the encoded display peptide and expression tag peptides.
Claims Coverage
The provided partial content identifies one independent claim. It recites a synthetic gene expression library with modified CDRH3s spanning about 4 to about 23 amino-acid residue lengths, expressed as first fusion proteins containing a display peptide and an expression tag, and featuring a defined repertoire size and specified human antibody variable heavy chain SEQ ID NO sets.
Synthetic gene expression library encoding first fusion proteins
A synthetic gene expression library encoding first fusion proteins, each comprising a fusion of a display peptide, a human antibody, and an expression tag, with a repertoire of 10^10 to 10^11 unique synthetic nucleic acid sequences encoding the first fusion proteins.
Modified CDRH3 with variable length from about 4 to about 23 amino-acid residues
Within each encoded variable heavy chain, the CDR3 region is modified from the respective SEQ ID NO, and the synthetic gene expression library encodes modified CDRH3s with lengths of 4 through 23 amino acid residues with amino-acid sequences defined by X1–Xn (SEQ ID NOs 113–132).
Human antibody variable heavy chain selected from specified SEQ ID NOs and corresponding nucleic acid sequences
Each encoded first fusion protein comprises a human antibody variable heavy chain with a polypeptide sequence selected from SEQ ID NOs. 2, 4, 6, 8, 10, 12 and 14 encoded by corresponding nucleic acid sequence selected from SEQ ID NOs. 1, 3, 5, 7, 9, 11 or 13.
Display peptide derived from a nucleotide sequence encoded in a display vector and expression tags selected from specified peptides
Each encoded first fusion protein includes a display peptide derived from a nucleotide sequence encoded in a display vector and at least one expression tag peptide selected from FLAG, c-Myc, (His)6-tag and V5-tag.
The identified independent claim centers on a large synthetic gene expression library encoding first fusion proteins that combine a display peptide and expression tag with a human antibody variable heavy chain bearing modified CDRH3s. The claim specifies the SEQ ID NO-based variable heavy chain selection and the modified CDRH3 design as defined by position-based X1–Xn sequences across CDRH3 lengths 4–23.
Stated Advantages
Not explicitly described in patent.
Documented Applications
Therapeutic, diagnostic, prognostic, and research uses are stated in the provided partial content.
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