Excision of HIV-1 DNA by gene editing: A proof-of-concept in vivo study

R. Kaminski, R. Bella, C. Yin, J. Otte, P. Ferrante, H. E. Gendelman, H. Li, R. Booze, J. Gordon, W. Hu, K. Khalili

Research output: Contribution to journalArticlepeer-review

154 Scopus citations


A CRISPR/Cas9 gene editing strategy has been remarkable in excising segments of integrated HIV-1 DNA sequences from the genome of latently infected human cell lines and by introducing InDel mutations, suppressing HIV-1 replication in patient-derived CD4+ T-cells, ex vivo. Here, we employed a short version of the Cas9 endonuclease, saCas9, together with a multiplex of guide RNAs (gRNAs) for targeting the viral DNA sequences within the 5'-LTR and the Gag gene for removing critically important segments of the viral DNA in transgenic mice and rats encompassing the HIV-1 genome. Tail-vein injection of transgenic mice with a recombinant Adeno-associated virus 9 (rAAV9) vector expressing saCas9 and the gRNAs, rAAV:saCas9/gRNA, resulted in the cleavage of integrated HIV-1 DNA and excision of a 978 bp DNA fragment spanning between the LTR and Gag gene in the spleen, liver, heart, lung and kidney as well as in the circulating lymphocytes. Retro-orbital inoculation of rAAV9:saCas9/gRNA in transgenic rats eliminated a targeted segment of viral DNA and substantially decreased the level of viral gene expression in circulating blood lymphocytes. The results from the proof-of-concept studies, for the first time, demonstrate the in vivo eradication of HIV-1 DNA by CRISPR/Cas9 on delivery by an rAAV9 vector in a range of cells and tissues that harbor integrated copies of viral DNA.

Original languageEnglish (US)
Pages (from-to)690-695
Number of pages6
JournalGene Therapy
Issue number8-9
StatePublished - Aug 1 2016

ASJC Scopus subject areas

  • Molecular Medicine
  • Molecular Biology
  • Genetics


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