Zinc finger nuclease-mediated CCR5 knockout hematopoietic stem cell transplantation controls HIV-1 in vivo

CCR5 is the major co-receptor used by HIV-1 and individuals homozygous for a 32bp deletion in CCR5 are profoundly resistant to HIV-1 infection. Using engineered zinc finger nucleases (ZFNs), we were able to disrupt the CCR5 gene in human hematopoietic stem/progenitor cells (HSC) at a mean frequency of 17% of total alleles in a population. This procedure produces both mono and bi-allelically disrupted cells. ZFN-treated HSC retained the ability to engraft NOD/SCID/IL2rγnull mice and gave rise to polyclonal multi-lineage progeny with the CCR5 gene permanently disrupted. Control mice receiving untreated HSC and challenged with CCR5-tropic HIV-1 displayed profound CD4+ T cell loss. In contrast, mice transplanted with ZFN-modified HSC underwent rapid selection for CCR5-negative cells, had significantly lower HIV-1 levels and preserved human cells throughout their tissues. The demonstration that a minority of CCR5-modified HSC can populate an infected animal with HIV-1-resistant, CCR5-negative progeny suggests the use of ZFN-modified autologous HSC as a clinical approach to treating HIV-1.