A novel CRISPR/Cas9-based nanomedicine strategy targeting CD4+ cells to cure HIV

Nearly 38 million people globally are living with HIV, the virus that causes AIDS by replicating in CD4+ immune cells. Current therapies can inhibit the virus from multiplying, but cannot eliminate viral DNA copies integrated in the genome of CD4+ long-living cells (viral reservoir). As such, HIV treatment requires HIV patients' lifelong commitment to drug adherence, which causes serious side effects, stigma, and a vast cost impact on society. An elegant approach to cure HIV would be to use gene therapy, but classical vectors remain unable to target the HIV reservoir. Here, we aim to design lipid nanoparticles (LNPs) coated with anti-CD4 nanobodies (Nb) that encode CRISPR/Cas9 ribonucleoproteins targeting both ends of the viral genome (long terminal repeats) and the viral coreceptor CCR5. As such, our novel nanomedicine will selectively purge HIV reservoirs in CD4+ cells, while conferring healthy CD4+ cells resistant to HIV reinfection in case of remnant virus. In addition, off-target effects in non-target cells will remain minimal. To do so, a LNP library along with different click-chemistry methods will be screened to obtain anti-CD4 Nb-coated LNPs with ideal pharmacophysical properties to encapsulate mRNA and transfect CD4+ cells. Selected LNPs will be further validated in vitro in primary human CD4+ cells and in vivo in a humanized mouse model of HIV infection. Together, the current proposal has a high potential to lay the foundation to make a final cure for HIV.