Optimization of a lentivirus-mediated gene therapy targeting HIV-1 RNA to eliminate HIV-1-infected cells
Year: 2024
Type: article
Abstract: Persistence of HIV-1 in cellular reservoirs results in lifelong infection, with cure achieved only in rare cases through ablation of marrow-derived cells. We report on optimization of an approach that could potentially be aimed at eliminating these reservoirs, hijacking the HIV-1 alternative splicing process to functionalize the herpes simplex virus thymidine kinase (HSVtk)/ganciclovir (GCV) cell suicide system through targeted RNA trans -splicing at the HIV-1 D4 donor site. AUG1-deficient HSVtk therapeutic pre-mRNA was designed to gain an in-frame start codon from HIV-1 tat1 . D4-targeting lentiviral vectors were produced and used to transduce HIV-1-expressing cells, where trans -spliced HIV-1 tat / HSVtk mRNA was successfully detected. However, translation of catalytically active HSVtk polypeptides from internal AUGs in HSVtk ΔAUG1 caused GCV-mediated cytotoxicity in uninfected cells. Modifying these sites in the D4 opt 2 lentiviral vector effectively mitigated this major off-target effect. Promoter choice was optimized for increased transgene expression. Affinity for HIV-1 RNA predicted in silico correlated with the propensity of opt 2 payloads to induce HIV-1 RNA trans -splicing and killing of HIV-1-expressing cells with no significant effect on uninfected cells. Following latency reversing agent (LRA) optimization and treatment, 45% of lymphocytes in an HIV-1-infected latency model could be eliminated with D4 opt 2/GCV. Further development would be warranted to exploit this approach.
Institutions: University of Cambridge, Addenbrooke's Hospital
Language: English
Subfield: Genetics
Domain: Life Sciences
Open Access status: gold
Estimated APC paid: $3,900
Funders: Higher Education Funding Council for England, Cancer Research UK, National Institute for Health and Care Research, University of Cambridge, Cambridge University Hospitals, National Institutes of Health, Medical Research Council, NIHR Cambridge Biomedical Research Centre, National Institute of Allergy and Infectious Diseases
OpenAlex ID: W4402564633