Single-hit genome edition for expression of single-chain immunoglobulins by edited B cells

Abstract

Lymphocytes have become attractive agents for adoptive immunotherapy but only the reformatting of T cells is efficiently mastered. Despite some recent breakthroughs, B cells remain challenging targets, with regard to both their long-term survival after in vitro manipulation and the rewiring of immunoglobulin (Ig) expression. Working on these two aspects, we have designed a new format of single-chain Ig ("scFull-Ig") coding cassette, the insertion of which at a single genomic position can redirect B cells toward a new antigen specificity, while preserving all functional domains of the B cell receptor. Precise genomic edition at a single locus then provides the most efficient and safe strategy to both disrupt endogenous Ig expression while encoding a new Ig paratope. As proofs of concept, functionality of such scFull BCR was validated by checking specific binding of two different classical targets of tumor immunotherapy, HER2 and CD20. Once the strategy validated in cell lines, it was also validated in primary B cells, again showing successful engineering of BCR expression. These results contribute to pave the way for future B cell-based adoptive cell therapy.