Selection of antibodies that regulate phenotype from intracellular combinatorial antibody libraries

Abstract

A method is presented that uses combinatorial antibody libraries to endow cells with new binding energy landscapes for the purpose of regulating their phenotypes. Antibodies that are expressed in cells infected with a lentiviral combinatorial antibody library are selected directly for function rather than only for binding. The potential diversity space can be very large because more than one lentivirus can infect a single cell. Thus, the initial combinatorial diversity of ∼1.0 × 10 11 members generated by the random association of antibody heavy and light chains is greatly increased by the reassortment of the antibody Fv domains themselves inside cells. The power of the system is illustrated by its ability to select unusual antibodies. Here, the selected antibodies are potent erythropoietin agonists whose ontogeny depends on recombination at the protein level of pairs of antibodies expressed in the same cell to generate heterodimeric bispecific antibodies. The obligate synergy between the different binding specificities of the antibody's monomeric subunits appears to replicate the asymmetric binding mechanism of authentic erythropoietin.agonist antibodies | phenotypic perturbation | erythropoietin phenocopy

Converting stem cells to dendritic cells by agonist antibodies from unbiased morphogenic selections

“…TF-1 erythroblast cells infected with the library were incorporated into methylcellulose agar for morphogenic screening. The antibody format that was used caused the antibodies to be secreted into the agar surrounding the cells (1). Compared with the control, the cells infected with members of the antibody library formed colonies with unique morphologies that included amorphous, scattered, linear, or round phenotypes (Fig.…”

“…This approach led to the identification of integrinbinding agonist antibodies that efficiently converted human stem cells to dendritic cells. intracellular combinatorial antibodies | phenotypic selections R ecently we developed combinatorial antibody libraries that are infectious for eukaryotic cells by incorporating them into lentiviruses (1). After infection, genotype and phenotype become linked because the antibody genes are incorporated into the genome and expressed and the cell, itself, becomes the reporter.…”

The Use of Antibodies in Small-Molecule Drug Discovery

“…A recent study has applied the intracellular expression of an antibody library in a eukaryotic cell using lentivirus, such that antibodies were expressed within the cell and secreted. 37 Using the erythropoetin receptor as proof of principle, a combination of expressed antibodies that gave potent erythropoietin agonism was identified. This work has been extended by the further use of antibody libraries in lentiviruses, infecting eukaryotic cells containing a fluorescent reporter coupled to a receptor.…”

Autocrine signaling based selection of combinatorial antibodies that transdifferentiate human stem cells

“…This method enables direct selection of agonists that regulate cellular phenotypes from the very cells expressing the agonist antibodies: i.e., in an autocrine manner (1). The power of these infectious libraries stems from several unique features.…”