Efficient editing of the CXCR4 locus using Cas9 ribonucleoprotein complexes stabilized with polyglutamic acid

Abstract

Gene editing using the CRISPR/Cas9 system provides new opportunities for the treatment of human diseases. Therefore, it is relevant to develop approaches aimed at increasing the efficiency of genome editing. Here, to increase the level of editing of the CXCR4 locus, a target for gene therapy of HIV infection, the Cas9 protein was modified by introducing additional NLS signals, and the ribonucleoprotein complexes of Cas9 and guide RNA were stabilized with poly-L-glutamic acid. This allowed a 1.8-fold increase in the level of CXCR4 knockout in the CEM/R5 T cell line and a 2-fold increase in the level of knock-in of the HIV-1 fusion peptide inhibitor MT-C34 in primary CD4+ T lymphocytes.