Fully human antagonistic antibodies against CCR4 potently inhibit cell signaling and chemotaxis.

Background: CC chemokine receptor 4 (CCR4) represents a potentially important target for cancer immunotherapy due to its expression on tumor infiltrating immune cells including regulatory T cells (Tregs) and on tumor cells in several cancer types and its role in metastasis.

Methodology: Using phage display, human antibody library, affinity maturation and a cell-based antibody selection strategy, the antibody variants against human CCR4 were generated. These antibodies effectively competed with ligand binding, were able to block ligand-induced signaling and cell migration, and demonstrated efficient killing of CCR4-positive tumor cells via ADCC and phagocytosis.

Screening human antibody libraries against carcinoma cells by affinity purification and polymerase chain reaction.

Bacterial scFv clones from a naïve antibody library have been isolated against cancer cell antigens with AffiSelect, a novel screening method that indirectly identifies candidate library members via an antigen reporter gene. The first step is the coating of carcinoma cell surface epitopes (antigen) with either mAbs, scFvs or phages (library members). Upon binding to a cell surface ligand, the library member generates a linking moiety.

Covalent antibody display--an in vitro antibody-DNA library selection system.

The endonuclease P2A initiates the DNA replication of the bacteriophage P2 by making a covalent bond with its own phosphate backbone. This enzyme has now been exploited as a new in vitro display tool for antibody fragments. We have constructed genetic fusions of P2A with single-chain antibodies (scFvs).