Selection of antibodies that regulate phenotype from intracellular combinatorial antibody libraries

Area of Science:

• Biotechnology
• Molecular Biology
• Immunology

Background:

• Engineering cellular functions requires precise control over molecular interactions.
• Traditional antibody selection often focuses on binding affinity, not functional output.
• Combinatorial antibody libraries offer vast potential for discovering novel protein functionalities.

Purpose of the Study:

• To develop a method for selecting functional antibodies directly from large combinatorial libraries.
• To engineer cells with novel binding properties for phenotype regulation.
• To demonstrate the system's capability in discovering unique antibodies with therapeutic potential.

Main Methods:

• Utilizing lentiviral combinatorial antibody libraries for intracellular expression.
• Employing a selection strategy based on antibody function, not just binding.
• Leveraging reassortment of antibody Fv domains within cells to expand diversity.
• Generating heterodimeric bispecific antibodies through protein-level recombination.

Main Results:

• Successfully selected potent erythropoietin agonists.
• Demonstrated the generation of bispecific antibodies via in-cell recombination.
• Showcased the ability to create antibodies with synergistic binding properties.
• Replicated the asymmetric binding mechanism of authentic erythropoietin.

Conclusions:

• The presented method enables direct functional selection of antibodies from diverse libraries.
• Intracellular antibody recombination significantly expands the accessible sequence space.
• This approach is powerful for discovering novel antibodies, including bispecific ones, with tailored functions.