An expanded genetic code facilitates antibody chemical conjugation involving the lambda light chain

Area of Science:

• Biotechnology
• Immunology
• Chemical Biology

Background:

• Current therapeutic antibodies predominantly use the immunoglobulin gamma (IgG) κ isotype.
• The IgGλ isotype presents a largely untapped resource for therapeutic antibody development.
• Chemical conjugation offers precise manufacturing of antibody conjugates, but sites in IgGλ remain underexplored.

Purpose of the Study:

• To explore efficient chemical conjugation strategies for the IgGλ isotype using an expanded genetic code.
• To identify specific conjugation sites within the IgGλ light chain constant domain (LCλ).
• To demonstrate the utility of site-specific conjugation for creating bispecific antibodies.

Main Methods:

• Incorporation of the unnatural amino acid Nε-(o-azidobenzyloxycarbonyl)-l-lysine (o-Az-Z-Lys) into the antigen-binding fragment (Fab) of IgGλ antibodies.
• Utilizing bio-orthogonal azido chemistry for site-specific chemical conjugation.
• Generating bispecific Fab-dimers by chemically linking modified cixutumumab and trastuzumab.

Main Results:

• Ten sites within the LCλ constant domain were identified as suitable for efficient chemical conjugation.
• These conjugation sites are largely specific to the λ isotype, differing from κ isotype sequences.
• The resulting bispecific Fab-dimers exhibited potent antagonistic activity against a cancer cell line.

Conclusions:

• The study successfully expands site-specific chemical conjugation methods to the IgGλ isotype.
• This approach enables precise modification and the creation of novel bispecific antibodies for therapeutic applications.
• The findings broaden the scope of chemical conjugation for the entire spectrum of humanized antibodies.