Once infected with HIV-1, the host immune system is incapable of ridding itself of the virus. HIV-1 uses latent viral reservoirs (LR) within CD4+ T cells to replicate. Within these reservoirs, HIV-1 is able to go into a latent state where it cannot be detected by the host's immune system or current HIV-1 treatments. By utilizing viewing assays and CRISPR-Cas9, there may be a possibility to identify, isolate, and then cut out HIV-1 from an infected cell. Two possible viewing assays have been proposed and studied in recent research. PCR assays are quicker and easier to administer while viral outgrowth assays (VOA) can measure the activation of the resting memory CD4+ T cells and the release of the HIV-1 from latency. Although research has shown that similar results may be able to be found from PCR, VOA is still the primary assay for HIV-1 LR studies. Current limitations of these assays include the PCR tending to overestimate the size of the LR and VOA underestimates the size, which, in turn, gives a possible false HIV-1 negative reading. Being able to view these LR at a precise reading is vital for the success of CRISPR-Cas9. Several strategies for the use of CRISPR-Cas9 have been proposed to aid in the excision of the HIV-1 LR from the genome of a cell, including saCas9 and spCas9. saCas9 would create a double-stranded break within the HIV-1 genetic code and would fully terminate the virus. Current research is focused on finding a viral transporter for the CRISPR-Cas9 that is big enough to fit the genetic coding needed to ensure the correct cleavage around the HIV-1 yet small enough to be successful. spCas9 is researched as a genome-editing tool that can cleave a eukaryotic double helix due to its double-stranded binding site. Along with dCas9, this technique can be used to potentially ‘cut out' the HIV-1 LRs and catalyze an HIV-1 reactivation which will cause virus-producing cell death. This ability causes both strands of the studied eukaryotic cell (in this case HIV-1) to be inactive and can thus repress the expression of the targeted genes. A method of delivery is needed to be studied in order for this strategy to be feasible.
Loechler, Sydney, "CRISPR/Cas9 Driven Eradication of HIV-1 in Infected Human Genome" (2019). Senior Seminars and Capstones. 1.