Stem cells, ribozymes and HIV

Abstract

In a recent Phase 2 clinical trial, Mitsuyasu et al. demonstrated that the delivery of an anti-HIV ribozyme from a gammaretroviral vector integrated into hematopoietic stem cells (HSCs) offers a safe and promising gene therapy strategy for the treatment of this infectious disease. This gene therapy strategy relies on the idea that, after the ex vivo transduction of an anti-HIV gene into autologous CD34+ HSCs, these human immunodeficiency virus (HIV)-resistant progenitors can be re-infused into an HIV patient, where they will differentiate and expand to permanently protect against the onset of AIDS (Figure 1). Although the inhibition of HIV has been considered a feasible and attractive candidate for gene therapy applications for over 20 years, this is the first reported randomized, double-blind cell-delivered Phase 2 clinical trial for HIV-1. The Mitsuyasu et al. study monitored 74 HIV-1-infected individuals for 100 weeks after infusion of transduced or control autologous HSCs. Thirty-eight of these patients were treated with HSCs carrying an integrated copy of a therapeutic, replication-incompetent, Moloney murine leukemia virus-based gammaretroviral vector (LNL6). The therapeutic vector (OZ1) expresses a hammerhead ribozyme against overlapping reading frames of the viral genes vpr and tat in unspliced and spliced viral transcripts, respectively. Although the researchers observed no significant differences in the viral load between OZ1 and control groups at any particular time point, they reported that transduced CD34+ cell therapy did not cause any apparent adverse effects and is safe for further exploration, thereby opening the doors for further trials with other gene therapy agents. Over the course of the 100-week clinical study, Mitsuyasu et al. observed sharp declines in the percentages of patients who maintained detectable levels of integrated DNA and mRNA from the OZ1 retroviral vector. In fact, at 20 weeks post-infusion, primary blood mononuclear cells from more than half of the treated patients had no detectable levels of OZ1 DNA or mRNA. This result is unexpected under the assumption that the OZ1-transduced HSCs successfully engrafted, differentiated and expanded into the T-cell lineage, and maintained some level of anti-HIV protection over untreated CD4+ T cells. Similar Phase I trials—including reports from the same authors—have confirmed successful engraftment, differentiation and expansion of autologous HSCs transduced with anti-HIV retroviral vectors. Therefore, although the HSCs may have failed to engraft under the conditions of the trial, it is also possible that the anti-HIV ribozyme failed to protect the OZ1 cells from HIV-1 infection and replication. This could be the result of mutational escape of the virus, suboptimal function of the ribozyme in vivo or possible shortcomings associated with gammaretroviral vectors. This Phase 2 investigation marks an important step toward a safe, cell-delivered gene therapy for HIV. However, to further advance this promising gene therapy approach, it will be necessary to identify the particular limitations and modify the existing therapeutic strategy. First, the virus might have acquired mutations in the target region of the ribozyme that would abrogate the function of the anti-HIV agent. Mutations arising at the second or third positions of the three-nucleotide ribozyme cleavage site of spliced and unspliced viral transcripts might render the anti-HIV ribozyme ineffective. Close examination of the targeted region suggests