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The Comprehensive NeuroAIDS Center (CNAC) Basic Science Core I: Gene editing strategy for eliminating HIV-1 DNA from cells.

This core now provides consultations on the use of gene editing strategy employing CRISPR/Cas9 for editing the gene of your interest in cell cultures and animal models. This service is provided by Dr. Rafal Kaminski who can be contacted at More recently, this technology has been utilized by Dr. Khalili’s laboratories for eradicating the HIV-1 genome from the human cell cultures, as described below.

Highly active antiretroviral therapy (HAART) profoundly inhibits HIV-1 replication, yet has failed to eliminate the virus from infected individuals. Thus, there is an urgent need for the development of effective strategies that will lead to the eradication of HIV-1 infection from the human population and the cure of AIDS. The presence of the proviral DNA genome as integrated copies in the host chromosomes, persistent low level replication of HIV-1 in latently infected cells, and continued risk for resurgence of the virus in resting CD4+ memory T-cells from deep dormancy leave little doubt that removal of the viral genome from the host chromosome is the ultimate cure for AIDS. Recently, we have successfully employed newly developed gene editing techniques, based on RNA-guided Cas9, to specifically target the HIV-1 genome and eliminate integrated copies of the proviral DNA from latently infected cell lines. In this attempt, we identified several specific DNA sequences within the U3 region of the HIV-1 LTR that serve as targets and are efficiently edited by single and multiplex Cas9/gRNAs to completely abrogate viral replication in latently infected macrophages, T-cells, and microglial cells. Our Cas9/gRNAs inflict no genotoxicity or off-target editing to the host cells, yet precisely excise a 9709 bp DNA fragment of the integrated proviral genome that spans between the 5'- and 3'- LTR. Moreover, the presence of multiplex gRNAs and Cas9 in the cells protected cells against future or subsequent HIV-1 infection. These observations offer a strong rationale to hypothesize that this novel approach can serve as a platform to explore the permanent removal of HIV-1 from the host and a cure for AIDS.