Unmet Need
Approximately 38 million people are currently living with human immunodeficiency virus (HIV) globally. HIV is a serious health issue, and around 1 million people die from HIV/AIDS related illnesses each year. Currently, there is no cure for HIV, only prevention and treatment techniques exist. Current methods of treatment use antiretroviral therapy (ART), which can suppress HIV, but cannot cure HIV. The virus is very difficult to eradicate due to the permanent integration of HIV genomes into the host cell chromosomes of immune cells, causing a latent pro-viral reservoir to be formed. Thus, there is a need for a way to specifically study and reverse HIV latency to pave the path to creation of targets and drugs that can help eradicate the disease.
Technology Overview
The inventors have constructed a Cas9-expressing cell line to serve as a polyclonal in vitro model for HIV-1 latency-related studies. The cell line consists of Jurkat-based cells, which were transduced by a GFP expressing defective lentiviral vector generated by the Karn Lab at Case Western University. The cells that exhibited GFP signals (GFP+) were sorted 5 days post-infection and were kept in culture for five weeks. Then, the modified post-sorting Jurkat cells were transduced by lentiCas9-Blast virus and turned into Cas9-expressing HIV-infected Jurkat cells. At the end of the 5-week incubation, cells without GFP expression (GFP-) were sorted, followed by TNF-⍺ stimulation. The GFP+ cells were sorted after the stimulation and kept in culture for another week. Finally, the GFP- cells were sorted and used as the latently HIV-infected polyclonal Jurkat cells for CRISPR screening. All GFP-based cell isolations were achieved at >99.9% purity. The inventors showed that HIV-1 gene expression can be re-induced in >90% of this cell population by TNF-⍺ treatment, as evidenced by GFP expression.
Stage of Development
The cell line is ready to use for the purposes of studying HIV-1 latency and developing latency reversal agents in academic and industrial settings.
