Human hematopoietic stem/progenitor cells modified by zinc-finger nucleases targeted to CCR5 control HIV-1 in vivo

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CCR5 is the major HIV-1 co-receptor, and individuals homozygous for a 32-bp deletion in CCR5 are resistant to infection by CCR5-tropic HIV-1. Using engineered zinc-finger nucleases (ZFNs), we disrupted CCR5 in human CD34+ hematopoietic stem/progenitor cells (HSPCs) at a mean frequency of 17% of the total alleles in a population. This procedure produces both mono- and bi-allelically disrupted cells. ZFN-treated HSPCs retained the ability to engraft NOD/SCID/IL2rγnull mice and gave rise to polyclonal multi-lineage progeny in which CCR5 was permanently disrupted. Control mice receiving untreated HSPCs and challenged with CCR5-tropic HIV-1 showed profound CD4+ T-cell loss. In contrast, mice transplanted with ZFN-modified HSPCs underwent rapid selection for CCR5−/− cells, had significantly lower HIV-1 levels and preserved human cells throughout their tissues. The demonstration that a minority of CCR5−/− HSPCs can populate an infected animal with HIV-1-resistant, CCR5−/− progeny supports the use of ZFN-modified autologous hematopoietic stem cells as a clinical approach to treating HIV-1.

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Figure 1: ZFN-mediated disruption of CCR5 in CD34+ HSPCs.
Figure 2: Protection of human CD4+ T cells in peripheral blood of HIV-infected mice previously engrafted with ZFN-modified CD34+ HSPCs.
Figure 3: Effects of HIV-1 infection on human cells in HSPC-engrafted NSG mice.
Figure 4: HIV-1 infection selects for disrupted CCR5 alleles.
Figure 5: ZFN activity produces heterogeneous mutations in CCR5.
Figure 6: Control of HIV-1 replication in mice receiving ZFN-treated CD34+ HSPCs.

Change history

  • 20 July 2010

    In the version of this article initially published online, the callout to Figure 6b was written incorrectly as Figure 6c. Also, in Figure 2b, the label on the y axis was missing a “/” between CD4+ and CD8+. The errors have been corrected for the print, PDF and HTML versions of this article.


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We would like to thank A. Cuddihy, S. Ge, R. Hollis and N. Smiley for expert technical assistance; C. Lutzko, V. Garcia, R. Akkina, B. Torbett and M. McCune for advice regarding humanized mice; and M. McCune for communicating unpublished data. This work was supported by funding from the California HIV/AIDS Research Project (P.M.C.), The Saban Research Institute (V.T.), and the National Heart, Lung, and Blood Institute P01 HL73104 (G.M.C., D.B.K. and P.M.C.).

Author information

N.H. performed most of the experiments; J.W., K.K., G.F. and X.W. developed assays and analyzed samples; V.T. contributed to discussions; N.H., G.M.C., D.B.K., P.D.G., M.C.H. and P.M.C. designed the experiments and analyzed data; N.H. and P.M.C. wrote the manuscript.

Correspondence to Paula M Cannon.

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J.W., K.K., G.F., P.D.G. and M.C.H. are employees of Sangamo BioSciences.

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