Preclinical safety and efficacy of an anti–HIV-1 lentiviral vector containing a short hairpin RNA to CCR5 and the C46 fusion inhibitor

Abstract

Gene transfer has therapeutic potential for treating HIV-1 infection by generating cells that are resistant to the virus. We have engineered a novel self-inactivating lentiviral vector, LVsh5/C46, using two viral-entry inhibitors to block early steps of HIV-1 cycle. The LVsh5/C46 vector encodes a short hairpin RNA (shRNA) for downregulation of CCR5, in combination with the HIV-1 fusion inhibitor, C46. We demonstrate here the effective delivery of LVsh5/C46 to human T cell lines, peripheral blood mononuclear cells, primary CD4+ T lymphocytes, and CD34+ hematopoietic stem/progenitor cells (HSPC). CCR5-targeted shRNA (sh5) and C46 peptide were stably expressed in the target cells and were able to effectively protect gene-modified cells against infection with CCR5- and CXCR4-tropic strains of HIV-1. LVsh5/C46 treatment was nontoxic as assessed by cell growth and viability, was noninflammatory, and had no adverse effect on HSPC differentiation. LVsh5/C46 could be produced at a scale sufficient for clinical development and resulted in active viral particles with very low mutagenic potential and the absence of replication-competent lentivirus. Based on these in vitro results, plus additional in vivo safety and efficacy data, LVsh5/C46 is now being tested in a phase 1/2 clinical trial for the treatment of HIV-1 disease.

Resolving the Sites of HIV Entry: Dynamin Networks Hold the Key

“…In an alternative approach, gene therapy can be used to deliver genes into target cells that can either protect them from new infections, by targeting viral entry [99][100][101] or inducing permanent silencing of the integrated provirus [102,103]. Deleting integrated provirus with gene editing systems such as CRISPR/Cas9 and zinc finger nucleases is also being explored [104][105][106][107][108].…”

Recent advances in RNAi-based strategies for therapy and prevention of HIV-1/AIDS

“…In particular, shRNA delivery via lentiviral vectors holds great promise, because shRNA continues to be produced following vector integration into the host genome, and thus a single treatment will have lasting effect. The concept here is that the modified cells, even at low-to-moderate levels of gene modification, have a selective survival advantage over unmodified cells, allowing them to proliferate and expand over time and, as a consequence, minimize viral loads and reduce viral reservoirs [129]. In fact, based on mathematical modeling it has been estimated that gene marking of ~10–20% in HSPCs would be sufficient to have an impact on viral load and CD4 + lymphocyte counts [130].…”

Stem cell-based therapies for HIV/AIDS

“…Importantly, rhesus macaque sh1005 modified CD4+ T cells were resistant to simian immunodeficiency virus (SIV) infection ex vivo [28], details of which have been described in the non-human primate section of this review (section 5.3). More recently, human sh1005 was combined with a second shRNA against HIV-LTR [32] or C46 HIV fusion inhibitor [33] to provide dual anti-HIV genes to inhibit both CCR5 tropic and CXCR4 tropic HIVs.…”

“…Therefore, it is possible that HIV may escape from C46, yet, this needs to be examined in vivo animal experiments. To minimize the emergence of HIV escape mutations, a dual anti-HIV combinatorial lentiviral vector for C46 CCR5 shRNA (sh1005) expression has been developed [33,60]. This vector is currently under investigation in a Phase I/II anti-HIV HSPC gene therapy clinical trial (NCT01734850, NCT02390297).…”

“…Vector transduced MOLT4/CCR5 and primary PBMC were resistant to both X4 and R5 tropic HIVs. (NL4-3 and BaL, respectively) [33]. The same vector combining sh1005 and C46 was then used to modify CD34+ HSPC and successfully reconstitute humanized BLT mice that supported multilineage human hematopoietic reconstitution.…”